Team:Lethbridge/Modeling
From 2010.igem.org
Liszabruder (Talk | contribs) |
Liszabruder (Talk | contribs) |
||
Line 125: | Line 125: | ||
=<font color="white">Homology Modeling= | =<font color="white">Homology Modeling= | ||
+ | |||
+ | Another aspect of our project is working on the localization of catechol-2,3,-dioxygenase (and other proteins) into the interior of <html><a href="https://2010.igem.org/Team:Lethbridge/Project/Compartamentalization"><font color="green"> microcompartments</font></a></html>. To target the protein into the microcompartment (Lumazine modified to have an even more charged interior) requires the fusion of a polyarginine tail to either the C or N terminus of the protein. | ||
+ | <br> | ||
+ | <br> | ||
+ | As with any fusion protien, the addition of this polyarginine tail to the protein has the very real potential of blocking (at least partially) the active site of a protein. Thus if one can predict the structure of the protein with the addition of the tail, it is possible to avoid creating inactive/ low efficiency fusion proteins. | ||
+ | |||
<html> | <html> | ||
<body> | <body> | ||
- | + | <center> | |
- | <table border="0" cellpadding="8" width="28%" style="background-color:#000000 | + | <table border="0" cellpadding="8" width="28%" style="background-color:#000000"> |
<tr> | <tr> | ||
<th> | <th> | ||
- | <img src="https://static.igem.org/mediawiki/2010/6/6f/Homology1.png" width=" | + | <img src="https://static.igem.org/mediawiki/2010/6/6f/Homology1.png" width="400"/> |
</th> | </th> | ||
</tr> | </tr> | ||
Line 139: | Line 145: | ||
<tr><th colspan="1" align="left"> | <tr><th colspan="1" align="left"> | ||
<p> | <p> | ||
- | <font color="white">Figure 1 | + | <font color="white">Figure 1. Homology model of xylE from <i>Pseudomonas putida</i>. The original structure (1MPY) is shown in teal; the generated homology model is shown in green. An Fe<sup>2+</sup> ion near the proposed active site is shown is brown. |
</th> | </th> | ||
</tr> | </tr> | ||
- | |||
</table> | </table> | ||
+ | </center> | ||
</body> | </body> | ||
</html> | </html> | ||
- | + | <br> | |
+ | ==<font color="white">The Method and Results== | ||
- | + | To model the xylE structure, the sequence for xylE from <i>Pseudomonas putida</i> (NCBI accession number NP_542866) was aligned with the primary sequence from the crystal structure of xylE from the same organism (pdb: 1MPY; several differences in amino acid sequence were observed) using CLUSTALW (Higgins <i>et al.</i>, 1996). Based on this sequence alignment, a homology model was generated using the alignment mode in SWISSMODEL (Guex <i>et al.</i>, 1997; Kiefer <i>et al.</i>, 2009; Arnold <i>et al.</i>, 2006). To model the placement of an N-terminal arginine tag, the tag was manually added to the N-terminus of the model. Energy minimization was carried out in SWISS-PDB viewer in vacuo utilizing a GROMOS96 energy minimization (Guex <i>et al.</i>, 1997). The resulting pdb file was visualized and manipulated using PYMOL, images were taken using the same software (DeLano, 2006). | |
- | + | ||
- | + | ||
- | |||
<html> | <html> | ||
<body> | <body> | ||
- | + | <center> | |
- | <table border="0" cellpadding="8" width="28%" style="background-color:#000000 | + | <table border="0" cellpadding="8" width="28%" style="background-color:#000000"> |
<tr> | <tr> | ||
<th> | <th> | ||
- | <img src="https://static.igem.org/mediawiki/2010/ | + | <img src="https://static.igem.org/mediawiki/2010/9/9b/Homology2.png" width="400"/> |
</th> | </th> | ||
</tr> | </tr> | ||
Line 167: | Line 171: | ||
<tr><th colspan="1" align="left"> | <tr><th colspan="1" align="left"> | ||
<p> | <p> | ||
- | <font color="white">Figure 2 | + | <font color="white">Figure 2. Energy minimized position of the N-terminal 10X Arg tag (shown in blue). |
</th> | </th> | ||
</tr> | </tr> | ||
</table> | </table> | ||
+ | </center> | ||
</body> | </body> | ||
</html> | </html> | ||
- | + | <br> | |
- | + | ||
=<font color="white">References= | =<font color="white">References= | ||
Arnold, K., Bordoli, L., Kopp, J., and Schwede, T. (2006) The SWISS-MODEL Workspace: A web-based environment for protein structure homology modelling., Bioinformatics 22, 195-201. | Arnold, K., Bordoli, L., Kopp, J., and Schwede, T. (2006) The SWISS-MODEL Workspace: A web-based environment for protein structure homology modelling., Bioinformatics 22, 195-201. |