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Página principal > Artículos > Artículos publicados > Crystal structure and mechanism of human carboxypeptidase O : |
Fecha: | 2018 |
Resumen: | Human metallocarboxypeptidase O (hCPO) is a recently discovered digestive enzyme localized to the apical membrane of intestinal epithelial cells. Unlike pancreatic metallocarboxypeptidases, hCPO is glycosylated and produced as an active enzyme with distinctive substrate specificity toward C-terminal (C-t) acidic residues. Here we present the crystal structure of hCPO at 1. 85-Å resolution, both alone and in complex with a carboxypeptidase inhibitor (NvCI) from the marine snail Nerita versicolor. The structure provides detailed information regarding determinants of enzyme specificity, in particular Arg275, placed at the bottom of the substrate-binding pocket. This residue, located at "canonical" position 255, where it is Ile in human pancreatic carboxypeptidases A1 (hCPA1) and A2 (hCPA2) and Asp in B (hCPB), plays a dominant role in determining the preference of hCPO for acidic C-t residues. Site-directed mutagenesis to Asp and Ala changes the specificity to C-t basic and hydrophobic residues, respectively. The single-site mutants thus faithfully mimic the enzymatic properties of CPB and CPA, respectively. hCPO also shows a preference for Glu over Asp, probably as a consequence of a tighter fitting of the Glu side chain in its S1' substrate-binding pocket. This unique preference of hCPO, together with hCPA1, hCPA2, and hCPB, completes the array of C-t cleavages enabling the digestion of the dietary proteins within the intestine. Finally, in addition to activity toward small synthetic substrates and peptides, hCPO can also trim C-t extensions of proteins, such as epidermal growth factor, suggesting a role in the maturation and degradation of growth factors and bioactive peptides. |
Ayudas: | Ministerio de Economía y Competitividad BFU2015-66417-P Ministerio de Economía y Competitividad BIO2016-78057-R Ministerio de Economía y Competitividad BES-2011-044872 Ministerio de Economía y Competitividad FPU12/06137 |
Derechos: | Tots els drets reservats. |
Lengua: | Anglès |
Documento: | Article ; recerca ; Versió acceptada per publicar |
Materia: | Carboxypeptidase ; Protein digestion ; Crystal structure ; Acidic protease |
Publicado en: | Proceedings of the National Academy of Sciences of the United States of America, Vol. 115, Issue 17 (April 2018) , p. E3932-E3939, ISSN 1091-6490 |
Postprint 29 p, 862.1 KB |