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Strongly hydrophobic proteins

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.

In order to provide our Bacillus subtilis biofilm with hydrophobic properties we have turned to highly hydrophobic proteins originating from Streptomyces coelicolor. This is a soil dwelling bacterium which undergoes morphological differentiation as it goes through different stages in its life cycle, greatly resembling fungal growth. After submerged, vegetative growth aerial hyphae are formed which protrude from the moist substrate. The formation of these aerial hyphae appear to require strongly hydrophobic proteins called chaplins, which have previously been described by Claessen et al (2003) and Elliot et al (2003).


Ant strepto.png
Ants strepto.png
Nature did it before! Tropical ants have a coating of streptomyces 
coelicolor on their exoskeleton, that produces antifungal sustances 
to protect them from pathogenic fungi.

Two subgroups

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.

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.

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.

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.

Physical properties

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.

Water and oil ten minutes after shaking, the separation of the two phases is clearly visable.

Besides hydrophobicity and hydrophilicity we came across even another very interesting property. When in monomeric form purified chaplins appear to have very good oil dispersing properties. When in emulsion with water in oil, chaplins will interact with both water and oil dispersing the oil into the water.

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.

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.

Overall these small proteins appear to have great potential in all kinds of applicational fields, making them very interesting additions to the parts registry as biobricks. However it was their most remarkable property – their hydrophobicity – which drew our attention to them to use them in our project.


Claessen, D; Rink, R; de Jong, W et al. 2003. A novel class of secreted hydrophobic proteins is involved in aerial hyphae formation in Streptomyces coelicolor by forming amyloid-like fibrils. Genes Dev 17 1714-1726

Elliot, MA; Karoonuthaisiri, N; Huang, JQ; et al. 2003 The chaplins: a family of hydrophobic cell-surface proteins involved in aerial mycelium formation in Streptomyces coelicolor. Genes Dev 17 1727-1740