Collagen prolyl 4-hydroxylases have sequence specificity towards different X-Pro-Gly triplets

Salo, A. M.; Rappu, P.; Koski, M. K.; Karjalainen, E.; Izzi, V.; Drushinin, K.; Miinalainen, I.; Käpylä, J.; Heino, J.; Myllyharju, J.

doi:10.1101/2023.06.28.546674

bioRxiv · 10.1101/2023.06.28.546674

Collagen prolyl 4-hydroxylases have sequence specificity towards different X-Pro-Gly triplets

Salo, A. M.·Rappu, P.·Koski, M. K.·Karjalainen, E.·Izzi, V.·Drushinin, K.·Miinalainen, I.·Käpylä, J.·Heino, J.·Myllyharju, J.

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Abstract

Formation of 4-hydroxyproline (4Hyp) in -X-Pro-Gly- collagen sequences is essential for the thermal stability of collagen molecules. 4Hyp formation is catalyzed by collagen prolyl 4-hydroxylases (C- P4H). Here we identify specific roles for the two main C-P4H isoenzymes by 4Hyp analysis of type I and IV collagens. Loss of C-P4H-I mainly affected prolines preceded by an X-position amino acid with a positively charged or a polar uncharged side chain. In contrast, loss of C-P4H-II affected triplets with a negatively charged glutamate or aspartate in the X-position, and their hydroxylation was found to be important as loss of C-P4H-II alone resulted in reduced collagen melting temperature and altered assembly of collagen fibrils and basement membrane. The C-P4H isoenzyme differences in substrate specificity were explained by selective substrate binding to the active site resulting in differences in Km and Vmax values. In conclusion, this study provides a molecular level explanation for the need of multiple C-P4H isoenzymes to generate collagen molecules capable to assemble into intact extracellular matrix structures.

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Salo, A. M., Rappu, P., Koski, M. K., Karjalainen, E., Izzi, V., Drushinin, K., Miinalainen, I., Käpylä, J., Heino, J., Myllyharju, J.. 2023-06-28. Collagen prolyl 4-hydroxylases have sequence specificity towards different X-Pro-Gly triplets. https://doi.org/10.1101/2023.06.28.546674

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