http://2010.igem.org/wiki/index.php?title=Team:Groningen/Hydrophobins&feed=atom&action=historyTeam:Groningen/Hydrophobins - Revision history2024-03-28T22:15:37ZRevision history for this page on the wikiMediaWiki 1.16.5http://2010.igem.org/wiki/index.php?title=Team:Groningen/Hydrophobins&diff=204305&oldid=prevMJvdN: /* Chaplins */2010-10-28T01:41:02Z<p><span class="autocomment">Chaplins</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Different chaplins, different functions ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Different chaplins, different functions ===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>There are a number of different chaplins shown in the figure below. In ''S. coelicolor'' these have specific functions in aerial growth and the transition to this phase. During submerged growth chaplins E and H are excreted and assemble at the water-air surface, drastically decreasing surface tension, allowing hyphae to break through the surface. On these forming aerial hyphae chaplins A to H assemble to form an extremely hydrophobic surface(Claessen ''et al'' 2003).</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>There are a number of different chaplins shown in the figure below. In ''S. coelicolor'' these have specific functions in aerial growth and the transition to this phase. During submerged growth chaplins E and H are excreted and assemble at the water-air surface, drastically decreasing surface tension, allowing hyphae to break through the surface. On these forming aerial hyphae chaplins A to H assemble to form an extremely hydrophobic surface(Claessen ''et al'' 2003).</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Groningen-ChaplinsStreptomcysis.png|center|600px|Showing the role of different chaplins during transition to aerial growth. Adapted from Claessen et al (2003).]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:Groningen-ChaplinsStreptomcysis.png|center|600px|Showing the role of different chaplins during transition to aerial growth. Adapted from Claessen et al (2003).]]</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><td></html>[[Image:Dispersant3GR.jpg|thumb|150px|Water and oil mixed with monomeric chaplins ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout the water phase.]]<html></td></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><td></html>[[Image:Dispersant3GR.jpg|thumb|150px|Water and oil mixed with monomeric chaplins ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout the water phase.]]<html></td></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><td></html>[[Image:Dispersant4GR.jpg|thumb|150px|Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.]]<html></td></div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div><td></html>[[Image:Dispersant4GR.jpg|thumb|150px|Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.]]<html></td></div></td></tr>
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</table>MJvdNhttp://2010.igem.org/wiki/index.php?title=Team:Groningen/Hydrophobins&diff=204255&oldid=prevMJvdN: /* Chaplins */2010-10-28T01:34:25Z<p><span class="autocomment">Chaplins</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Chaplins ==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>== Chaplins ==</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''Summary'''</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''Summary'''</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Introduction ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Introduction ===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We do not just want to make a biological coating, we would also give it special properties. We focused our attention on surface hydrophobicity since this might lead to very interesting applications. One remarkable example of surface hydrophobicity can be found in nature on [http://www.youtube.com/watch?v=MFHcSrNRU5E lotus leafs], which are extremely water repellant. Interestingly, these leafs are self-cleansing due to this property. Surface hydrophobicity has overall been shown to have self-cleansing and anti-fouling properties (Nimittrakoolchai ''et al'' 2007) and is subject of much research. However, to obtain hydrophobic surface activity mostly chemical techniques are used. We sought to use a biological alternative.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We do not just want to make a biological coating, we would also give it special properties. We focused our attention on surface hydrophobicity since this might lead to very interesting applications. One remarkable example of surface hydrophobicity can be found in nature on [http://www.youtube.com/watch?v=MFHcSrNRU5E lotus leafs], which are extremely water repellant. Interestingly, these leafs are self-cleansing due to this property. Surface hydrophobicity has overall been shown to have self-cleansing and anti-fouling properties (Nimittrakoolchai ''et al'' 2007) and is subject of much research. However, to obtain hydrophobic surface activity mostly chemical techniques are used. We sought to use a biological alternative.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Two subgroups ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Two subgroups ===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Chaplins can be categorized into two groups. The first group consists of chaplins A to C and are about 225 amino acids in size. These large chaplins contain a signal sequence, two hydrophobic chaplin domains, a hydrophilic region and a cell wall anchor. The second group includes chaplin D to H and are with around 63 amino acids smaller than the afore mentioned chaplins. Being smaller, they only contain a signal sequence followed by a hydrophobic chaplin domain.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Chaplins can be categorized into two groups. The first group consists of chaplins A to C and are about 225 amino acids in size. These large chaplins contain a signal sequence, two hydrophobic chaplin domains, a hydrophilic region and a cell wall anchor. The second group includes chaplin D to H and are with around 63 amino acids smaller than the afore mentioned chaplins. Being smaller, they only contain a signal sequence followed by a hydrophobic chaplin domain.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:ChpCartoon.jpg|center|600px|The hydrophobic chaplin domains are shown in green and are present on all chaplins. The hydrophillic region and cell wall anchor of the large chaplins are shown in blue and red, respectively.]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:ChpCartoon.jpg|center|600px|The hydrophobic chaplin domains are shown in green and are present on all chaplins. The hydrophillic region and cell wall anchor of the large chaplins are shown in blue and red, respectively.]]</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Chaplins have the intrinsic property of assembling into rod-like structures called amyloid fibers. These fibers are very rigid and hard to break down and even resist boiling in SDS which denatures almost all natural occuring proteins. They share distinguishing features with the medically important amyloid fibers that are characteristic for many neurodegenerative diseases such as Alzheimer's, Huntington's, systemic amyloidosis and the prion diseases.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Chaplins have the intrinsic property of assembling into rod-like structures called amyloid fibers. These fibers are very rigid and hard to break down and even resist boiling in SDS which denatures almost all natural occuring proteins. They share distinguishing features with the medically important amyloid fibers that are characteristic for many neurodegenerative diseases such as Alzheimer's, Huntington's, systemic amyloidosis and the prion diseases.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Moreover, chaplins are extremely stable, both thermally and chemically. As an illustration, to purify them one has to turn to severe techniques like boiling in SDS and extraction with trifluoroacetic acid. Also, along the entire duration of our project – more than half a year – we did not observe any decline in the physical properties of our purified chaplins, being able to re-use the proteins over and over again.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Moreover, chaplins are extremely stable, both thermally and chemically. As an illustration, to purify them one has to turn to severe techniques like boiling in SDS and extraction with trifluoroacetic acid. Also, along the entire duration of our project – more than half a year – we did not observe any decline in the physical properties of our purified chaplins, being able to re-use the proteins over and over again.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Chaplins in our project ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Chaplins in our project ===</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We chose not to make all chaplins into biobricks, but to focus on chaplin C, E and H. Chaplin C is anchored in the cell wall and together with chaplins E and H coat the outside of the cell with a hydrophobic layer. Before ordering our biobricks we codon optimalized them for ''B. subtilis''. Since these proteins are not native to ''B. subtilis'' and we were unsure whether it would not degrade them with proteases we tested chaplin degradation in spent medium of ''B. subtilis''. Degradation appeared not to occur. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We chose not to make all chaplins into biobricks, but to focus on chaplin C, E and H. Chaplin C is anchored in the cell wall and together with chaplins E and H coat the outside of the cell with a hydrophobic layer. Before ordering our biobricks we codon optimalized them for ''B. subtilis''. Since these proteins are not native to ''B. subtilis'' and we were unsure whether it would not degrade them with proteases we tested chaplin degradation in spent medium of ''B. subtilis''. Degradation appeared not to occur. </div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:ChpBB.jpg|center|600px|A cartoon showing our chaplin biobricks. In addition to the regular C chaplin we added one which has an sortase binding site (shown in yellow).]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:ChpBB.jpg|center|600px|A cartoon showing our chaplin biobricks. In addition to the regular C chaplin we added one which has an sortase binding site (shown in yellow).]]</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, we did not know whether ''B. subtilis'' would successfully secrete the C chaplin and insert it into its cell wall. So in addition to chaplins [http://partsregistry.org/wiki/index.php?title=Part:BBa_K305001 C], [http://partsregistry.org/wiki/index.php?title=Part:BBa_K305003 E] and [http://partsregistry.org/wiki/index.php?title=Part:BBa_K305004 H] we also designed an [http://partsregistry.org/wiki/index.php?title=Part:BBa_K305002 altered C chaplin] which has an added sortase binding motive and included a [http://partsregistry.org/wiki/index.php?title=Part:BBa_K305005 sortase] BioBrick to our project. Sortase - originating from ''Staphylococcus aureus'' - is a well studied protein which anchors surface proteins to the cell wal (Mazmanian ''et al'' 1999).</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, we did not know whether ''B. subtilis'' would successfully secrete the C chaplin and insert it into its cell wall. So in addition to chaplins [http://partsregistry.org/wiki/index.php?title=Part:BBa_K305001 C], [http://partsregistry.org/wiki/index.php?title=Part:BBa_K305003 E] and [http://partsregistry.org/wiki/index.php?title=Part:BBa_K305004 H] we also designed an [http://partsregistry.org/wiki/index.php?title=Part:BBa_K305002 altered C chaplin] which has an added sortase binding motive and included a [http://partsregistry.org/wiki/index.php?title=Part:BBa_K305005 sortase] BioBrick to our project. Sortase - originating from ''Staphylococcus aureus'' - is a well studied protein which anchors surface proteins to the cell wal (Mazmanian ''et al'' 1999).</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Physical properties ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Physical properties ===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:chaplins.jpg|<del class="diffchange diffchange-inline">left</del>|300px|The normally hydrophobic surface of the petri dish on the left is made hydrophillic by coating it with purified chaplins as seen on the right<del class="diffchange diffchange-inline">.]][[Image:Dispersant4GR.jpg|thumb|150px|right|Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.]][[Image:Dispersant3GR.jpg|thumb|150px|right|Water and oil mixed with monomeric chaplins ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout the water phase</del>.]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:chaplins.jpg|<ins class="diffchange diffchange-inline">right</ins>|300px|The normally hydrophobic surface of the petri dish on the left is made hydrophillic by coating it with purified chaplins as seen on the right.]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Interestingly, purified chaplins can be used to coat normally hydrophobic surfaces such as petri dishes, rendering them hydrophilic. This is due to their amphipatic nature, being hydrophobic on one side and hydrophilic on the other. In nature though they only coat the outside of the aerial hyphae of ''S. coelicolor'' hydrophobically. We pose that the assembly on the outside of cells is important for the amyloid fibers to polymerize into the right configuration to obtain extreme hydrophobicity. This is one of the reasons we chose to express chaplins in a biofilm as opposed to coat surfaces with purified chaplins. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Interestingly, purified chaplins can be used to coat normally hydrophobic surfaces such as petri dishes, rendering them hydrophilic. This is due to their amphipatic nature, being hydrophobic on one side and hydrophilic on the other. In nature though they only coat the outside of the aerial hyphae of ''S. coelicolor'' hydrophobically. We pose that the assembly on the outside of cells is important for the amyloid fibers to polymerize into the right configuration to obtain extreme hydrophobicity. This is one of the reasons we chose to express chaplins in a biofilm as opposed to coat surfaces with purified chaplins. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del style="color: red; font-weight: bold; text-decoration: none;">[[Image:Dispersant1GR.jpg|thumb|300px|center|Left to right: Water and oil; Water, oil, Congo Red staining and AssembledChaplins; Water, oil, Congo Red staining and monomeric chaplins; Water, oil and Congo Red staining.]]</del></div></td><td colspan="2"> </td></tr>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"><html></ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"><table></ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"><tr></ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"><td></html>[[Image:Dispersant3GR.jpg|thumb|150px|Water and oil mixed with monomeric chaplins ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout the water phase.]]<html></td></ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"><td></html>[[Image:Dispersant1GR.jpg|thumb|300px|Left to right: Water and oil; Water, oil, Congo Red staining and AssembledChaplins; Water, oil, Congo Red staining and monomeric chaplins; Water, oil and Congo Red staining.]] <html></td></ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"><td></html>[[Image:Dispersant4GR.jpg|thumb|150px|Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.]]<html></td></ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"></tr></ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"></table></ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;"></html></ins></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Moreover, chaplins are extremely stable, both thermally and chemically. As an illustration, to purify them one has to turn to severe techniques like boiling in SDS and extraction with trifluoroacetic acid. Also, along the entire duration of our project – more than half a year – we did not observe any decline in the physical properties of our purified chaplins, being able to re-use the proteins over and over again.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Moreover, chaplins are extremely stable, both thermally and chemically. As an illustration, to purify them one has to turn to severe techniques like boiling in SDS and extraction with trifluoroacetic acid. Also, along the entire duration of our project – more than half a year – we did not observe any decline in the physical properties of our purified chaplins, being able to re-use the proteins over and over again.</div></td></tr>
</table>MJvdNhttp://2010.igem.org/wiki/index.php?title=Team:Groningen/Hydrophobins&diff=203932&oldid=prevMJvdN: /* Chaplins */2010-10-28T01:20:04Z<p><span class="autocomment">Chaplins</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We do not just want to make a biological coating, we would also give it special properties. We focused our attention on surface hydrophobicity since this might lead to very interesting applications. One remarkable example of surface hydrophobicity can be found in nature on [http://www.youtube.com/watch?v=MFHcSrNRU5E lotus leafs], which are extremely water repellant. Interestingly, these leafs are self-cleansing due to this property. Surface hydrophobicity has overall been shown to have self-cleansing and anti-fouling properties (Nimittrakoolchai ''et al'' 2007) and is subject of much research. However, to obtain hydrophobic surface activity mostly chemical techniques are used. We sought to use a biological alternative.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We do not just want to make a biological coating, we would also give it special properties. We focused our attention on surface hydrophobicity since this might lead to very interesting applications. One remarkable example of surface hydrophobicity can be found in nature on [http://www.youtube.com/watch?v=MFHcSrNRU5E lotus leafs], which are extremely water repellant. Interestingly, these leafs are self-cleansing due to this property. Surface hydrophobicity has overall been shown to have self-cleansing and anti-fouling properties (Nimittrakoolchai ''et al'' 2007) and is subject of much research. However, to obtain hydrophobic surface activity mostly chemical techniques are used. We sought to use a biological alternative.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:ChpMicro.gif|right|200px|Electron microscopy picture showing S. coelicolor spores on which chaplins can be seen clearly as rod-like structures. Cover of Journal of Bacteriology, September 2008.]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:ChpMicro.gif<ins class="diffchange diffchange-inline">|thumb</ins>|right|200px|Electron microscopy picture showing S. coelicolor spores on which chaplins can be seen clearly as rod-like structures. Cover of Journal of Bacteriology, September 2008.]]</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Physical properties ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Physical properties ===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:chaplins.jpg|left|300px|The normally hydrophobic surface of the petri dish on the left is made hydrophillic by coating it with purified chaplins as seen on the right.]][[Image:Dispersant4GR.jpg|150px|right|Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.]][[Image:Dispersant3GR.jpg|150px|right|Water and oil mixed with monomeric chaplins ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout the water phase.]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:chaplins.jpg|left|300px|The normally hydrophobic surface of the petri dish on the left is made hydrophillic by coating it with purified chaplins as seen on the right.]][[Image:Dispersant4GR.jpg<ins class="diffchange diffchange-inline">|thumb</ins>|150px|right|Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.]][[Image:Dispersant3GR.jpg<ins class="diffchange diffchange-inline">|thumb</ins>|150px|right|Water and oil mixed with monomeric chaplins ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout the water phase.]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Interestingly, purified chaplins can be used to coat normally hydrophobic surfaces such as petri dishes, rendering them hydrophilic. This is due to their amphipatic nature, being hydrophobic on one side and hydrophilic on the other. In nature though they only coat the outside of the aerial hyphae of ''S. coelicolor'' hydrophobically. We pose that the assembly on the outside of cells is important for the amyloid fibers to polymerize into the right configuration to obtain extreme hydrophobicity. This is one of the reasons we chose to express chaplins in a biofilm as opposed to coat surfaces with purified chaplins. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Interestingly, purified chaplins can be used to coat normally hydrophobic surfaces such as petri dishes, rendering them hydrophilic. This is due to their amphipatic nature, being hydrophobic on one side and hydrophilic on the other. In nature though they only coat the outside of the aerial hyphae of ''S. coelicolor'' hydrophobically. We pose that the assembly on the outside of cells is important for the amyloid fibers to polymerize into the right configuration to obtain extreme hydrophobicity. This is one of the reasons we chose to express chaplins in a biofilm as opposed to coat surfaces with purified chaplins. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:Dispersant1GR.jpg|300px|center|Left to right: Water and oil; Water, oil, Congo Red staining and AssembledChaplins; Water, oil, Congo Red staining and monomeric chaplins; Water, oil and Congo Red staining.]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:Dispersant1GR.jpg<ins class="diffchange diffchange-inline">|thumb</ins>|300px|center|Left to right: Water and oil; Water, oil, Congo Red staining and AssembledChaplins; Water, oil, Congo Red staining and monomeric chaplins; Water, oil and Congo Red staining.]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
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</table>MJvdNhttp://2010.igem.org/wiki/index.php?title=Team:Groningen/Hydrophobins&diff=203196&oldid=prevMJvdN at 00:58, 28 October 20102010-10-28T00:58:41Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:ChpBB.jpg|center|600px|A cartoon showing our chaplin biobricks. In addition to the regular C chaplin we added one which has an sortase binding site (shown in yellow).]]</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[Image:ChpBB.jpg|center|600px|A cartoon showing our chaplin biobricks. In addition to the regular C chaplin we added one which has an sortase binding site (shown in yellow).]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Also, we did not know whether ''B. subtilis'' would successfully secrete the C chaplin and insert it into its cell wall. So in addition to chaplins C, E and H we also designed an altered C chaplin which has an added sortase binding motive and included a sortase <del class="diffchange diffchange-inline">biobrick </del>to our project. Sortase - originating from ''Staphylococcus aureus'' - is a well studied protein which anchors surface proteins to the cell wal (Mazmanian ''et al'' 1999).</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Also, we did not know whether ''B. subtilis'' would successfully secrete the C chaplin and insert it into its cell wall. So in addition to chaplins <ins class="diffchange diffchange-inline">[http://partsregistry.org/wiki/index.php?title=Part:BBa_K305001 </ins>C<ins class="diffchange diffchange-inline">]</ins>, <ins class="diffchange diffchange-inline">[http://partsregistry.org/wiki/index.php?title=Part:BBa_K305003 </ins>E<ins class="diffchange diffchange-inline">] </ins>and <ins class="diffchange diffchange-inline">[http://partsregistry.org/wiki/index.php?title=Part:BBa_K305004 </ins>H<ins class="diffchange diffchange-inline">] </ins>we also designed an <ins class="diffchange diffchange-inline">[http://partsregistry.org/wiki/index.php?title=Part:BBa_K305002 </ins>altered C chaplin<ins class="diffchange diffchange-inline">] </ins>which has an added sortase binding motive and included a <ins class="diffchange diffchange-inline">[http://partsregistry.org/wiki/index.php?title=Part:BBa_K305005 </ins>sortase<ins class="diffchange diffchange-inline">] BioBrick </ins>to our project. Sortase - originating from ''Staphylococcus aureus'' - is a well studied protein which anchors surface proteins to the cell wal (Mazmanian ''et al'' 1999).</div></td></tr>
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</table>MJvdNhttp://2010.igem.org/wiki/index.php?title=Team:Groningen/Hydrophobins&diff=200207&oldid=prevMJvdN: /* Chaplins */2010-10-27T23:13:51Z<p><span class="autocomment">Chaplins</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We do not just want to make a biological coating, we would also give it special properties. We focused our attention on surface hydrophobicity since this might lead to very interesting applications. One remarkable example of surface hydrophobicity can be found in nature on [http://www.youtube.com/watch?v=MFHcSrNRU5E lotus leafs], which are extremely water repellant. Interestingly, these leafs are self-cleansing due to this property. Surface hydrophobicity has overall been shown to have self-cleansing and anti-fouling properties (Nimittrakoolchai ''et al'' 2007) and is subject of much research. However, to obtain hydrophobic surface activity mostly chemical techniques are used. We sought to use a biological alternative.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We do not just want to make a biological coating, we would also give it special properties. We focused our attention on surface hydrophobicity since this might lead to very interesting applications. One remarkable example of surface hydrophobicity can be found in nature on [http://www.youtube.com/watch?v=MFHcSrNRU5E lotus leafs], which are extremely water repellant. Interestingly, these leafs are self-cleansing due to this property. Surface hydrophobicity has overall been shown to have self-cleansing and anti-fouling properties (Nimittrakoolchai ''et al'' 2007) and is subject of much research. However, to obtain hydrophobic surface activity mostly chemical techniques are used. We sought to use a biological alternative.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:ChpMicro.gif<del class="diffchange diffchange-inline">|thumb</del>|right|200px|Electron microscopy picture showing S. coelicolor spores on which chaplins can be seen clearly as rod-like structures. Cover of Journal of Bacteriology, September 2008.]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:ChpMicro.gif|right|200px|Electron microscopy picture showing S. coelicolor spores on which chaplins can be seen clearly as rod-like structures. Cover of Journal of Bacteriology, September 2008.]]</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>There are a number of different chaplins shown in the figure below. In ''S. coelicolor'' these have specific functions in aerial growth and the transition to this phase. During submerged growth chaplins E and H are excreted and assemble at the water-air surface, drastically decreasing surface tension, allowing hyphae to break through the surface. On these forming aerial hyphae chaplins A to H assemble to form an extremely hydrophobic surface(Claessen ''et al'' 2003).</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>There are a number of different chaplins shown in the figure below. In ''S. coelicolor'' these have specific functions in aerial growth and the transition to this phase. During submerged growth chaplins E and H are excreted and assemble at the water-air surface, drastically decreasing surface tension, allowing hyphae to break through the surface. On these forming aerial hyphae chaplins A to H assemble to form an extremely hydrophobic surface(Claessen ''et al'' 2003).</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:Groningen-ChaplinsStreptomcysis.png<del class="diffchange diffchange-inline">|thumb</del>|center|600px|Showing the role of different chaplins during transition to aerial growth. Adapted from Claessen et al (2003).]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:Groningen-ChaplinsStreptomcysis.png|center|600px|Showing the role of different chaplins during transition to aerial growth. Adapted from Claessen et al (2003).]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Two subgroups ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Two subgroups ===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Chaplins can be categorized into two groups. The first group consists of chaplins A to C and are about 225 amino acids in size. These large chaplins contain a signal sequence, two hydrophobic chaplin domains, a hydrophilic region and a cell wall anchor. The second group includes chaplin D to H and are with around 63 amino acids smaller than the afore mentioned chaplins. Being smaller, they only contain a signal sequence followed by a hydrophobic chaplin domain.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Chaplins can be categorized into two groups. The first group consists of chaplins A to C and are about 225 amino acids in size. These large chaplins contain a signal sequence, two hydrophobic chaplin domains, a hydrophilic region and a cell wall anchor. The second group includes chaplin D to H and are with around 63 amino acids smaller than the afore mentioned chaplins. Being smaller, they only contain a signal sequence followed by a hydrophobic chaplin domain.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:ChpCartoon.jpg<del class="diffchange diffchange-inline">|thumb</del>|center|600px|The hydrophobic chaplin domains are shown in green and are present on all chaplins. The hydrophillic region and cell wall anchor of the large chaplins are shown in blue and red, respectively.]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:ChpCartoon.jpg|center|600px|The hydrophobic chaplin domains are shown in green and are present on all chaplins. The hydrophillic region and cell wall anchor of the large chaplins are shown in blue and red, respectively.]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Chaplins have the intrinsic property of assembling into rod-like structures called amyloid fibers. These fibers are very rigid and hard to break down and even resist boiling in SDS which denatures almost all natural occuring proteins. They share distinguishing features with the medically important amyloid fibers that are characteristic for many neurodegenerative diseases such as Alzheimer's, Huntington's, systemic amyloidosis and the prion diseases.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Chaplins have the intrinsic property of assembling into rod-like structures called amyloid fibers. These fibers are very rigid and hard to break down and even resist boiling in SDS which denatures almost all natural occuring proteins. They share distinguishing features with the medically important amyloid fibers that are characteristic for many neurodegenerative diseases such as Alzheimer's, Huntington's, systemic amyloidosis and the prion diseases.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Physical properties ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Physical properties ===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:chaplins.jpg<del class="diffchange diffchange-inline">|thumb</del>|left|300px|The normally hydrophobic surface of the petri dish on the left is made hydrophillic by coating it with purified chaplins as seen on the right.]][[Image:Dispersant4GR.jpg<del class="diffchange diffchange-inline">|thumb</del>|150px|right|Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.]][[Image:Dispersant3GR.jpg<del class="diffchange diffchange-inline">|thumb</del>|150px|right|Water and oil mixed with monomeric chaplins ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout the water phase.]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:chaplins.jpg|left|300px|The normally hydrophobic surface of the petri dish on the left is made hydrophillic by coating it with purified chaplins as seen on the right.]][[Image:Dispersant4GR.jpg|150px|right|Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.]][[Image:Dispersant3GR.jpg|150px|right|Water and oil mixed with monomeric chaplins ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout the water phase.]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Interestingly, purified chaplins can be used to coat normally hydrophobic surfaces such as petri dishes, rendering them hydrophilic. This is due to their amphipatic nature, being hydrophobic on one side and hydrophilic on the other. In nature though they only coat the outside of the aerial hyphae of ''S. coelicolor'' hydrophobically. We pose that the assembly on the outside of cells is important for the amyloid fibers to polymerize into the right configuration to obtain extreme hydrophobicity. This is one of the reasons we chose to express chaplins in a biofilm as opposed to coat surfaces with purified chaplins. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Interestingly, purified chaplins can be used to coat normally hydrophobic surfaces such as petri dishes, rendering them hydrophilic. This is due to their amphipatic nature, being hydrophobic on one side and hydrophilic on the other. In nature though they only coat the outside of the aerial hyphae of ''S. coelicolor'' hydrophobically. We pose that the assembly on the outside of cells is important for the amyloid fibers to polymerize into the right configuration to obtain extreme hydrophobicity. This is one of the reasons we chose to express chaplins in a biofilm as opposed to coat surfaces with purified chaplins. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:Dispersant1GR.jpg<del class="diffchange diffchange-inline">|thumb</del>|300px|center|Left to right: Water and oil; Water, oil, Congo Red staining and AssembledChaplins; Water, oil, Congo Red staining and monomeric chaplins; Water, oil and Congo Red staining.]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:Dispersant1GR.jpg|300px|center|Left to right: Water and oil; Water, oil, Congo Red staining and AssembledChaplins; Water, oil, Congo Red staining and monomeric chaplins; Water, oil and Congo Red staining.]]</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We chose not to make all chaplins into biobricks, but to focus on chaplin C, E and H. Chaplin C is anchored in the cell wall and together with chaplins E and H coat the outside of the cell with a hydrophobic layer. Before ordering our biobricks we codon optimalized them for ''B. subtilis''. Since these proteins are not native to ''B. subtilis'' and we were unsure whether it would not degrade them with proteases we tested chaplin degradation in spent medium of ''B. subtilis''. Degradation appeared not to occur. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We chose not to make all chaplins into biobricks, but to focus on chaplin C, E and H. Chaplin C is anchored in the cell wall and together with chaplins E and H coat the outside of the cell with a hydrophobic layer. Before ordering our biobricks we codon optimalized them for ''B. subtilis''. Since these proteins are not native to ''B. subtilis'' and we were unsure whether it would not degrade them with proteases we tested chaplin degradation in spent medium of ''B. subtilis''. Degradation appeared not to occur. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:ChpBB.jpg<del class="diffchange diffchange-inline">|thumb</del>|center|600px|A cartoon showing our chaplin biobricks. In addition to the regular C chaplin we added one which has an sortase binding site (shown in yellow).]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:ChpBB.jpg|center|600px|A cartoon showing our chaplin biobricks. In addition to the regular C chaplin we added one which has an sortase binding site (shown in yellow).]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, we did not know whether ''B. subtilis'' would successfully secrete the C chaplin and insert it into its cell wall. So in addition to chaplins C, E and H we also designed an altered C chaplin which has an added sortase binding motive and included a sortase biobrick to our project. Sortase - originating from ''Staphylococcus aureus'' - is a well studied protein which anchors surface proteins to the cell wal (Mazmanian ''et al'' 1999).</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, we did not know whether ''B. subtilis'' would successfully secrete the C chaplin and insert it into its cell wall. So in addition to chaplins C, E and H we also designed an altered C chaplin which has an added sortase binding motive and included a sortase biobrick to our project. Sortase - originating from ''Staphylococcus aureus'' - is a well studied protein which anchors surface proteins to the cell wal (Mazmanian ''et al'' 1999).</div></td></tr>
</table>MJvdNhttp://2010.igem.org/wiki/index.php?title=Team:Groningen/Hydrophobins&diff=199464&oldid=prevMJvdN: /* Physical properties */2010-10-27T22:54:22Z<p><span class="autocomment">Physical properties</span></p>
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<td colspan='2' style="background-color: white; color:black;">Revision as of 22:54, 27 October 2010</td>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Physical properties ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Physical properties ===</div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">[[Image:chaplins.jpg|thumb|left|300px|The normally hydrophobic surface of the petri dish on the left is made hydrophillic by coating it with purified chaplins as seen on the right.]][[Image:Dispersant4GR.jpg|thumb|150px|right|Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.]][[Image:Dispersant3GR.jpg|thumb|150px|right|Water and oil mixed with monomeric chaplins ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout the water phase.]]</ins></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Interestingly, purified chaplins can be used to coat normally hydrophobic surfaces such as petri dishes, rendering them hydrophilic. This is due to their amphipatic nature, being hydrophobic on one side and hydrophilic on the other. In nature though they only coat the outside of the aerial hyphae of ''S. coelicolor'' hydrophobically. We pose that the assembly on the outside of cells is important for the amyloid fibers to polymerize into the right configuration to obtain extreme hydrophobicity. This is one of the reasons we chose to express chaplins in a biofilm as opposed to coat surfaces with purified chaplins. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Interestingly, purified chaplins can be used to coat normally hydrophobic surfaces such as petri dishes, rendering them hydrophilic. This is due to their amphipatic nature, being hydrophobic on one side and hydrophilic on the other. In nature though they only coat the outside of the aerial hyphae of ''S. coelicolor'' hydrophobically. We pose that the assembly on the outside of cells is important for the amyloid fibers to polymerize into the right configuration to obtain extreme hydrophobicity. This is one of the reasons we chose to express chaplins in a biofilm as opposed to coat surfaces with purified chaplins. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">[[Image:Dispersant1GR.jpg|thumb|300px|center|Left to right: Water and oil; Water, oil, Congo Red staining and AssembledChaplins; Water, oil, Congo Red staining and monomeric chaplins; Water, oil and Congo Red staining.]]</ins></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del style="color: red; font-weight: bold; text-decoration: none;">Moreover, chaplins are extremely stable, both thermally and chemically. As an illustration, to purify them one has to turn to severe techniques like boiling in SDS and extraction with trifluoroacetic acid. Also, along the entire duration of our project – more than half a year – we did not observe any decline in the physical properties of our purified chaplins, being able to re-use the proteins over and over again.</del></div></td><td colspan="2"> </td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">[[Image:</del>chaplins<del class="diffchange diffchange-inline">.jpg|thumb|300px|The normally hydrophobic surface of the petri dish on the left is made hydrophillic by coating it with purified chaplins as seen on the right.]]</del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">Moreover, </ins>chaplins <ins class="diffchange diffchange-inline">are extremely stable</ins>, <ins class="diffchange diffchange-inline">both thermally </ins>and <ins class="diffchange diffchange-inline">chemically. As an illustration</ins>, <ins class="diffchange diffchange-inline">to purify them one has to turn to severe techniques like boiling in SDS </ins>and <ins class="diffchange diffchange-inline">extraction with trifluoroacetic acid</ins>. <ins class="diffchange diffchange-inline">Also</ins>, <ins class="diffchange diffchange-inline">along </ins>the <ins class="diffchange diffchange-inline">entire duration </ins>of <ins class="diffchange diffchange-inline">our project – more than half a year – we did not observe any decline in </ins>the <ins class="diffchange diffchange-inline">physical properties of our purified </ins>chaplins<ins class="diffchange diffchange-inline">, being able to re-use </ins>the <ins class="diffchange diffchange-inline">proteins over and over again</ins>.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">[[Image:Dispersant1GR.jpg|thumb|300px|Left to right: Water and oil; Water</del>, <del class="diffchange diffchange-inline">oil, Congo Red staining </del>and <del class="diffchange diffchange-inline">AssembledChaplins; Water</del>, <del class="diffchange diffchange-inline">oil, Congo Red staining </del>and <del class="diffchange diffchange-inline">monomeric chaplins; Water, oil and Congo Red staining</del>.<del class="diffchange diffchange-inline">]][[Image:Dispersant4GR.jpg|thumb|150px|Water and oil ten minutes after shaking</del>, the <del class="diffchange diffchange-inline">separation </del>of the <del class="diffchange diffchange-inline">two phases is clearly visable.]]</del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">[[Image:Dispersant3GR.jpg|thumb|150px|Water and oil mixed with monomeric </del>chaplins <del class="diffchange diffchange-inline">ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout </del>the <del class="diffchange diffchange-inline">water phase</del>.<del class="diffchange diffchange-inline">]]</del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div></div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Chaplins in our project ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Chaplins in our project ===</div></td></tr>
</table>MJvdNhttp://2010.igem.org/wiki/index.php?title=Team:Groningen/Hydrophobins&diff=199405&oldid=prevMJvdN: /* Physical properties */2010-10-27T22:52:50Z<p><span class="autocomment">Physical properties</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Physical properties ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Physical properties ===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del style="color: red; font-weight: bold; text-decoration: none;">[[Image:chaplins.jpg|thumb|left|300px|The normally hydrophobic surface of the petri dish on the left is made hydrophillic by coating it with purified chaplins as seen on the right.]][[Image:Dispersant4GR.jpg|thumb|150px|right|Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.]][[Image:Dispersant3GR.jpg|thumb|150px|right|Water and oil mixed with monomeric chaplins ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout the water phase.]]</del></div></td><td colspan="2"> </td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Interestingly, purified chaplins can be used to coat normally hydrophobic surfaces such as petri dishes, rendering them hydrophilic. This is due to their amphipatic nature, being hydrophobic on one side and hydrophilic on the other. In nature though they only coat the outside of the aerial hyphae of ''S. coelicolor'' hydrophobically. We pose that the assembly on the outside of cells is important for the amyloid fibers to polymerize into the right configuration to obtain extreme hydrophobicity. This is one of the reasons we chose to express chaplins in a biofilm as opposed to coat surfaces with purified chaplins. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Interestingly, purified chaplins can be used to coat normally hydrophobic surfaces such as petri dishes, rendering them hydrophilic. This is due to their amphipatic nature, being hydrophobic on one side and hydrophilic on the other. In nature though they only coat the outside of the aerial hyphae of ''S. coelicolor'' hydrophobically. We pose that the assembly on the outside of cells is important for the amyloid fibers to polymerize into the right configuration to obtain extreme hydrophobicity. This is one of the reasons we chose to express chaplins in a biofilm as opposed to coat surfaces with purified chaplins. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del style="color: red; font-weight: bold; text-decoration: none;">[[Image:Dispersant1GR.jpg|thumb|300px|center|Left to right: Water and oil; Water, oil, Congo Red staining and AssembledChaplins; Water, oil, Congo Red staining and monomeric chaplins; Water, oil and Congo Red staining.]]</del></div></td><td colspan="2"> </td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del style="color: red; font-weight: bold; text-decoration: none;"></del></div></td><td colspan="2"> </td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Moreover, chaplins are extremely stable, both thermally and chemically. As an illustration, to purify them one has to turn to severe techniques like boiling in SDS and extraction with trifluoroacetic acid. Also, along the entire duration of our project – more than half a year – we did not observe any decline in the physical properties of our purified chaplins, being able to re-use the proteins over and over again.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Moreover, chaplins are extremely stable, both thermally and chemically. As an illustration, to purify them one has to turn to severe techniques like boiling in SDS and extraction with trifluoroacetic acid. Also, along the entire duration of our project – more than half a year – we did not observe any decline in the physical properties of our purified chaplins, being able to re-use the proteins over and over again.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">[[Image:chaplins.jpg|thumb|300px|The normally hydrophobic surface of the petri dish on the left is made hydrophillic by coating it with purified chaplins as seen on the right.]]</ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">[[Image:Dispersant1GR.jpg|thumb|300px|Left to right: Water and oil; Water, oil, Congo Red staining and AssembledChaplins; Water, oil, Congo Red staining and monomeric chaplins; Water, oil and Congo Red staining.]][[Image:Dispersant4GR.jpg|thumb|150px|Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.]]</ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">[[Image:Dispersant3GR.jpg|thumb|150px|Water and oil mixed with monomeric chaplins ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout the water phase.]]</ins></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Chaplins in our project ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Chaplins in our project ===</div></td></tr>
</table>MJvdNhttp://2010.igem.org/wiki/index.php?title=Team:Groningen/Hydrophobins&diff=199213&oldid=prevMJvdN: /* Chaplins */2010-10-27T22:45:44Z<p><span class="autocomment">Chaplins</span></p>
<table style="background-color: white; color:black;">
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>There are a number of different chaplins shown in the figure below. In ''S. coelicolor'' these have specific functions in aerial growth and the transition to this phase. During submerged growth chaplins E and H are excreted and assemble at the water-air surface, drastically decreasing surface tension, allowing hyphae to break through the surface. On these forming aerial hyphae chaplins A to H assemble to form an extremely hydrophobic surface(Claessen ''et al'' 2003).</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>There are a number of different chaplins shown in the figure below. In ''S. coelicolor'' these have specific functions in aerial growth and the transition to this phase. During submerged growth chaplins E and H are excreted and assemble at the water-air surface, drastically decreasing surface tension, allowing hyphae to break through the surface. On these forming aerial hyphae chaplins A to H assemble to form an extremely hydrophobic surface(Claessen ''et al'' 2003).</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:Groningen-ChaplinsStreptomcysis.png|thumb|center|Showing the role of different chaplins during transition to aerial growth. Adapted from Claessen et al (2003).]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:Groningen-ChaplinsStreptomcysis.png|thumb|center<ins class="diffchange diffchange-inline">|600px</ins>|Showing the role of different chaplins during transition to aerial growth. Adapted from Claessen et al (2003).]]</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Two subgroups ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Two subgroups ===</div></td></tr>
</table>MJvdNhttp://2010.igem.org/wiki/index.php?title=Team:Groningen/Hydrophobins&diff=199180&oldid=prevMJvdN: /* Chaplins */2010-10-27T22:43:48Z<p><span class="autocomment">Chaplins</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We do not just want to make a biological coating, we would also give it special properties. We focused our attention on surface hydrophobicity since this might lead to very interesting applications. One remarkable example of surface hydrophobicity can be found in nature on [http://www.youtube.com/watch?v=MFHcSrNRU5E lotus leafs], which are extremely water repellant. Interestingly, these leafs are self-cleansing due to this property. Surface hydrophobicity has overall been shown to have self-cleansing and anti-fouling properties (Nimittrakoolchai ''et al'' 2007) and is subject of much research. However, to obtain hydrophobic surface activity mostly chemical techniques are used. We sought to use a biological alternative.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We do not just want to make a biological coating, we would also give it special properties. We focused our attention on surface hydrophobicity since this might lead to very interesting applications. One remarkable example of surface hydrophobicity can be found in nature on [http://www.youtube.com/watch?v=MFHcSrNRU5E lotus leafs], which are extremely water repellant. Interestingly, these leafs are self-cleansing due to this property. Surface hydrophobicity has overall been shown to have self-cleansing and anti-fouling properties (Nimittrakoolchai ''et al'' 2007) and is subject of much research. However, to obtain hydrophobic surface activity mostly chemical techniques are used. We sought to use a biological alternative.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:ChpMicro.gif|right|200px|Electron microscopy picture showing S. coelicolor spores on which chaplins can be seen clearly as rod-like structures. Cover of Journal of Bacteriology, September 2008.]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:ChpMicro.gif<ins class="diffchange diffchange-inline">|thumb</ins>|right|200px|Electron microscopy picture showing S. coelicolor spores on which chaplins can be seen clearly as rod-like structures. Cover of Journal of Bacteriology, September 2008.]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>There are a number of different chaplins shown in the figure below. In ''S. coelicolor'' these have specific functions in aerial growth and the transition to this phase. During submerged growth chaplins E and H are excreted and assemble at the water-air surface, drastically decreasing surface tension, allowing hyphae to break through the surface. On these forming aerial hyphae chaplins A to H assemble to form an extremely hydrophobic surface(Claessen ''et al'' 2003).</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>There are a number of different chaplins shown in the figure below. In ''S. coelicolor'' these have specific functions in aerial growth and the transition to this phase. During submerged growth chaplins E and H are excreted and assemble at the water-air surface, drastically decreasing surface tension, allowing hyphae to break through the surface. On these forming aerial hyphae chaplins A to H assemble to form an extremely hydrophobic surface(Claessen ''et al'' 2003).</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:Groningen-ChaplinsStreptomcysis.png|center|Showing the role of different chaplins during transition to aerial growth. Adapted from Claessen et al (2003).]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:Groningen-ChaplinsStreptomcysis.png<ins class="diffchange diffchange-inline">|thumb</ins>|center|Showing the role of different chaplins during transition to aerial growth. Adapted from Claessen et al (2003).]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Two subgroups ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Two subgroups ===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Chaplins can be categorized into two groups. The first group consists of chaplins A to C and are about 225 amino acids in size. These large chaplins contain a signal sequence, two hydrophobic chaplin domains, a hydrophilic region and a cell wall anchor. The second group includes chaplin D to H and are with around 63 amino acids smaller than the afore mentioned chaplins. Being smaller, they only contain a signal sequence followed by a hydrophobic chaplin domain.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Chaplins can be categorized into two groups. The first group consists of chaplins A to C and are about 225 amino acids in size. These large chaplins contain a signal sequence, two hydrophobic chaplin domains, a hydrophilic region and a cell wall anchor. The second group includes chaplin D to H and are with around 63 amino acids smaller than the afore mentioned chaplins. Being smaller, they only contain a signal sequence followed by a hydrophobic chaplin domain.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:ChpCartoon.jpg|center|600px|The hydrophobic chaplin domains are shown in green and are present on all chaplins. The hydrophillic region and cell wall anchor of the large chaplins are shown in blue and red, respectively.]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:ChpCartoon.jpg<ins class="diffchange diffchange-inline">|thumb</ins>|center|600px|The hydrophobic chaplin domains are shown in green and are present on all chaplins. The hydrophillic region and cell wall anchor of the large chaplins are shown in blue and red, respectively.]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Chaplins have the intrinsic property of assembling into rod-like structures called amyloid fibers. These fibers are very rigid and hard to break down and even resist boiling in SDS which denatures almost all natural occuring proteins. They share distinguishing features with the medically important amyloid fibers that are characteristic for many neurodegenerative diseases such as Alzheimer's, Huntington's, systemic amyloidosis and the prion diseases.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Chaplins have the intrinsic property of assembling into rod-like structures called amyloid fibers. These fibers are very rigid and hard to break down and even resist boiling in SDS which denatures almost all natural occuring proteins. They share distinguishing features with the medically important amyloid fibers that are characteristic for many neurodegenerative diseases such as Alzheimer's, Huntington's, systemic amyloidosis and the prion diseases.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Physical properties ===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=== Physical properties ===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:chaplins.jpg|left|300px|The normally hydrophobic surface of the petri dish on the left is made hydrophillic by coating it with purified chaplins as seen on the right.]][[Image:Dispersant4GR.jpg|150px|right|Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.]][[Image:Dispersant3GR.jpg|150px|right|Water and oil mixed with monomeric chaplins ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout the water phase.]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:chaplins.jpg<ins class="diffchange diffchange-inline">|thumb</ins>|left|300px|The normally hydrophobic surface of the petri dish on the left is made hydrophillic by coating it with purified chaplins as seen on the right.]][[Image:Dispersant4GR.jpg<ins class="diffchange diffchange-inline">|thumb</ins>|150px|right|Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.]][[Image:Dispersant3GR.jpg<ins class="diffchange diffchange-inline">|thumb</ins>|150px|right|Water and oil mixed with monomeric chaplins ten minutes after shaking. Chaplins are stained with Congo Red. The oil is clearly dispersed throughout the water phase.]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Interestingly, purified chaplins can be used to coat normally hydrophobic surfaces such as petri dishes, rendering them hydrophilic. This is due to their amphipatic nature, being hydrophobic on one side and hydrophilic on the other. In nature though they only coat the outside of the aerial hyphae of ''S. coelicolor'' hydrophobically. We pose that the assembly on the outside of cells is important for the amyloid fibers to polymerize into the right configuration to obtain extreme hydrophobicity. This is one of the reasons we chose to express chaplins in a biofilm as opposed to coat surfaces with purified chaplins. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Interestingly, purified chaplins can be used to coat normally hydrophobic surfaces such as petri dishes, rendering them hydrophilic. This is due to their amphipatic nature, being hydrophobic on one side and hydrophilic on the other. In nature though they only coat the outside of the aerial hyphae of ''S. coelicolor'' hydrophobically. We pose that the assembly on the outside of cells is important for the amyloid fibers to polymerize into the right configuration to obtain extreme hydrophobicity. This is one of the reasons we chose to express chaplins in a biofilm as opposed to coat surfaces with purified chaplins. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, when monomeric, chaplins show to reduce surface tension of water which is illustrated in the figure below. It is clearly visable in the second tube from the right that monomeric chaplins flatten the meniscus at the water-oil interface. However, once assembled into polymeres, this property is lost as can be seen in the second tube from the left, which shows the same features as the tube on the far left which only contains water and oil. The tube on the far right only contained water, oil and Congo Red staining.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:Dispersant1GR.jpg|300px|center|Left to right: Water and oil; Water, oil, Congo Red staining and AssembledChaplins; Water, oil, Congo Red staining and monomeric chaplins; Water, oil and Congo Red staining.]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:Dispersant1GR.jpg<ins class="diffchange diffchange-inline">|thumb</ins>|300px|center|Left to right: Water and oil; Water, oil, Congo Red staining and AssembledChaplins; Water, oil, Congo Red staining and monomeric chaplins; Water, oil and Congo Red staining.]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We chose not to make all chaplins into biobricks, but to focus on chaplin C, E and H. Chaplin C is anchored in the cell wall and together with chaplins E and H coat the outside of the cell with a hydrophobic layer. Before ordering our biobricks we codon optimalized them for ''B. subtilis''. Since these proteins are not native to ''B. subtilis'' and we were unsure whether it would not degrade them with proteases we tested chaplin degradation in spent medium of ''B. subtilis''. Degradation appeared not to occur. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We chose not to make all chaplins into biobricks, but to focus on chaplin C, E and H. Chaplin C is anchored in the cell wall and together with chaplins E and H coat the outside of the cell with a hydrophobic layer. Before ordering our biobricks we codon optimalized them for ''B. subtilis''. Since these proteins are not native to ''B. subtilis'' and we were unsure whether it would not degrade them with proteases we tested chaplin degradation in spent medium of ''B. subtilis''. Degradation appeared not to occur. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>[[Image:ChpBB.jpg|center|600px|A cartoon showing our chaplin biobricks. In addition to the regular C chaplin we added one which has an sortase binding site (shown in yellow).]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>[[Image:ChpBB.jpg<ins class="diffchange diffchange-inline">|thumb</ins>|center|600px|A cartoon showing our chaplin biobricks. In addition to the regular C chaplin we added one which has an sortase binding site (shown in yellow).]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, we did not know whether ''B. subtilis'' would successfully secrete the C chaplin and insert it into its cell wall. So in addition to chaplins C, E and H we also designed an altered C chaplin which has an added sortase binding motive and included a sortase biobrick to our project. Sortase - originating from ''Staphylococcus aureus'' - is a well studied protein which anchors surface proteins to the cell wal (Mazmanian ''et al'' 1999).</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Also, we did not know whether ''B. subtilis'' would successfully secrete the C chaplin and insert it into its cell wall. So in addition to chaplins C, E and H we also designed an altered C chaplin which has an added sortase binding motive and included a sortase biobrick to our project. Sortase - originating from ''Staphylococcus aureus'' - is a well studied protein which anchors surface proteins to the cell wal (Mazmanian ''et al'' 1999).</div></td></tr>
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