The Peptide-Substrate-binding Domain of Collagen Prolyl 4-Hydroxylases Is a Tetratricopeptide Repeat Domain with Functional Aromatic Residues

• Published in: The Journal of biological chemistry 2004-12, Vol.279 (50), p.52255-52261
• Main Authors: Pekkala, Mira, Hieta, Reija, Bergmann, Ulrich, Kivirikko, Kari I, Wierenga, Rik K, Myllyharju, Johanna
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Catalytic Domain; Collagen - biosynthesis; Crystallography, X-Ray; Humans; Hydrophobic and Hydrophilic Interactions; In Vitro Techniques; Models, Molecular; Molecular Sequence Data; Mutagenesis, Site-Directed; Procollagen-Proline Dioxygenase - chemistry; Procollagen-Proline Dioxygenase - genetics; Procollagen-Proline Dioxygenase - metabolism; Protein Structure, Tertiary; Protein Subunits; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Repetitive Sequences, Amino Acid; Substrate Specificity
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M410007200

Collagen prolyl 4-hydroxylases catalyze the formation of 4-hydroxyproline in - X -Pro-Gly-sequences and have an essential role in collagen synthesis. The vertebrate enzymes are α 2 β 2 tetramers in which the catalytic α-subunits contain separate peptide-substrate-binding and catalytic domains. We report on the crystal structure of the peptide-substrate-binding domain of the human type I enzyme refined at 2.3 Å resolution. It was found to belong to a family of tetratricopeptide repeat domains that are involved in many protein-protein interactions and consist of five α-helices forming two tetratricopeptide repeat motifs plus the solvating helix. A prominent feature of its concave surface is a deep groove lined by tyrosines, a putative binding site for proline-rich Tripeptides. Solvent-exposed side chains of three of the tyrosines have a repeat distance similar to that of a poly- l -proline type II helix. The aromatic surface ends at one of the tyrosines, where the groove curves almost 90° away from the linear arrangement of the three tyrosine side chains, possibly inducing a bent conformation in the bound peptide. This finding is consistent with previous suggestions by others that a minimal structural requirement for proline 4-hydroxylation may be a sequence in the poly- l -proline type II conformation followed by a β-turn in the Pro-Gly segment. Site-directed mutagenesis indicated that none of the tyrosines was critical for tetramer assembly, whereas most of them were critical for the binding of a peptide substrate and inhibitor both to the domain and the α 2 β 2 enzyme tetramer.