The P3H1 complex has disulfide isomerase activity in vitro 1 An Additional Function of the Rough Endoplasmic Reticulum Protein complex Prolyl 3- Hydroxylase 1∙Cartilage associated protein∙Cyclophilin B: The CXXXC motif Reveals Disulfide Isomerase Activity in vitro

Collagen biosynthesis occurs in the rough Endoplasmic Reticulum (rER) and many molecular chaperones and folding enzymes are involved in this process. The folding mechanism of type I procollagen has been well characterized and protein disulfide isomerase (PDI) has been suggested as a key player in the formation of the correct disulfide bonds in the non-collagenous carboxy-terminal and aminoterminal propeptides. Prolyl 3-hydroxylase 1 (P3H1) forms a hetero-trimeric complex with cartilage associated protein (CRTAP) and Cyclophilin B (CypB). This complex is a multi functional complex acting as a prolyl 3hydroxylase, a peptidyl prolyl cis-trans isomerase and a molecular chaperone. Two major domains are predicted from the primary sequence of P3H1: An amino-terminal domain and a carboxy-terminal domain corresponding to the 2-oxoglutarateand iron-dependant dioxygenase domains similar to the α-subunit of prolyl 4-hydroxylase and lysyl hydroxylases. The amino-terminal domain contains four CXXXC sequence repeats. The primary sequence of CRTAP is homologous to the amino-terminal domain of P3H1 and also contains four CXXXC sequence repeats. However, the function of the CXXXC sequence repeats is not known. Several publications have reported that short peptides containing a CXC or a CXXC sequence show oxido-reductase activity similar to PDI in vitro. We hypothesize that CXXXC motifs have oxido-reductase activity similar to the CXXC motif in PDI. We have tested the enzyme activities on model substrates in vitro using a GCRALCG peptide and the P3H1 complex. Our results suggest that this complex could function as a disulfide isomerase in the rER. _______________________________________