2026-04-17T17:00:44Zhttps://recercat.cat/oai/request

oai:recercat.cat:2072/4394812026-03-13T05:51:49Zcom_2072_98col_2072_378192

urn:hdl:2072/439481 Crystal structure and mechanism of human carboxypeptidase O : insights into its specific activity for acidic residues García Guerrero, María del Carmen Garcia-Pardo, Javier Berenguer De La Cuesta, Esther Fernandez-Alvarez, Roberto Barfi, Gifty B. Lyons, Peter J. Avilés, Francesc Xavier Huber, Robert Lorenzo Rivera, Julia Reverter Cendrós, David Carboxypeptidase Protein digestion Crystal structure Acidic protease 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. 2018 Article Ministerio de Economía y Competitividad BFU2015-66417-P Agencia Estatal de Investigación BIO2016-78057-R Ministerio de Ciencia e Innovación BES-2011-044872 Ministerio de Educación, Cultura y Deporte FPU12/06137 Proceedings of the National Academy of Sciences of the United States of America ; Vol. 115, Issue 17 (April 2018), p. E3932-E3939 open access Aquest material està protegit per drets d'autor i/o drets afins. Podeu utilitzar aquest material en funció del que permet la legislació de drets d'autor i drets afins d'aplicació al vostre cas. Per a d'altres usos heu d'obtenir permís del(s) titular(s) de drets. https://rightsstatements.org/vocab/InC/1.0/