Structure of transmembrane prolyl 4-hydroxylase reveals unique organization of EF and dioxygenase domains.
Prolyl 4-hydroxylases (P4Hs) catalyze post-translational hydroxylation of peptidyl proline residues. In addition to collagen P4Hs and hypoxia-inducible factor P4Hs, a third P4H-the poorly characterized endoplasmic reticulum-localized transmembrane prolyl 4-hydroxylase (P4H-TM)-is found in animals. P4H-TM variants are associated with the familiar neurological HIDEA syndrome, but how these variants might contribute to disease is unknown. Here, we explored this question in a structural and functional analysis of soluble human P4H-TM. The crystal structure revealed an EF domain with two Ca2+-binding motifs inserted within the catalytic domain. A substrate-binding groove was formed between the EF domain and the conserved core of the catalytic domain. The proximity of the EF domain to the active site suggests that Ca2+ binding is relevant to the catalytic activity. Functional analysis demonstrated that Ca2+-binding affinity of P4H-TM is within the range of physiological Ca2+ concentration in the endoplasmic reticulum. P4H-TM was found both as a monomer and a dimer in the solution, but the monomer-dimer equilibrium was not regulated by Ca2+. The catalytic site contained bound Fe2+ and N-oxalylglycine, which is an analogue of the cosubstrate 2-oxoglutarate. Comparison with homologous P4H structures complexed with peptide substrates showed that the substrate-interacting residues and the lid structure that folds over the substrate are conserved in P4H-TM, whereas the extensive loop structures that surround the substrate-binding groove, generating a negative surface potential, are different. Analysis of the structure suggests that the HIDEA variants cause loss of P4H-TM function. In conclusion, P4H-TM shares key structural elements with other P4Hs while having a unique EF domain.
['Myllykoski|Matti|M|', 'Sutinen|Aleksi|A|', 'Koski|M Kristian|MK|', 'Kallio|Juha P|JP|', 'Raasakka|Arne|A|', 'Myllyharju|Johanna|J|', 'Wierenga|Rik K|RK|', 'Koivunen|Peppi|P|']
D018360:Crystallography, X-Ray D049308:Dioxygenases D020832:EF Hand Motifs D006801:Humans D008958:Models, Molecular D064828:Prolyl Hydroxylases D011487:Protein Conformation D000072417:Protein Domains
prolyl 4-hydroxylase HIDEA syndrome endoplasmic reticulum 2OGDD EF hand calcium binding