Association of a bacterial collagen after hydroxyproline incorporation

Abstract

Various bacterial collagen-like proteins have been previously described and shown to have a triple helical, (Gly-Xaa Yaa)_n repeating structure. They are stable without needing any secondary modification of proline residues in the Yaa position to hydroxyproline, a characteristic feature of animal collagens. Hydroxyproline can, however, be introduced into recombinant bacterial collagen by co-translational incorporation during fermentation. However, this does not lead to full incorporation and introduces the hydroxyproline into both the Xaa and Yaa positions. It was suggested that the poor solubility of bacterial collagen samples with higher levels of incorporation of hydroxyproline could be due to an increase in protein association at neutral pH. In the present study, cryo-transmission electron microscopy was used to examine the nature and extent of any associations arising from hydroxyproline incorporation. This was examined further, using 2 smaller fragments, where the proline sites are predominantly in either the Xaa position or Yaa position. The present data confirm the importance of the presence of hydroxyproline in assisting in the association between collagen molecules.