Structure and Mechanism of Metallocarboxypeptidases

Abstract

Metallocarboxpeptidases cleave C-terminal residues from peptide substrates and participate in a wide range of physiological processes, but they also contribute to human pathology. On the basis of structural information, we can distinguish between two groups of such metallopeptidases: cowrins and funnelins. Cowrins comprise protozoan, prokaryotic, and mammalian enzymes related to both neurolysin and angiotensin-converting enzyme and their catalytic domains contain 500–700 residues. They are ellipsoidal and traversed horizontally by a long, deep, narrow active-site cleft, in which the C-terminal residues are cut from oligopeptides and unstructured protein tails. The consensus cowrin structure contains a common core of 17 helices and a three-stranded β-sheet, which participates in substrate binding. This protease family is characterized by a set of spatially conserved amino acids involved in catalysis, HEXXH+EXXS/G+H+Y/R+Y. Funnelins comprise structural relatives of the archetypal bovine carboxypeptidase A1 and feature mammalian, insect and bacterial proteins with strict carboxypeptidase activity. Their ∼ 300-residue catalytic domains evince a consensus central eight-stranded β-sheet flanked on either side by a total of eight helices. They also contain a characteristic set of conserved residues, HXXE+R+NR+H+Y+E, and their active-site clefts are rather shallow and lie at the bottom of a funnel-like cavity. Therefore, these enzymes act on a large variety of well-folded proteins. In both cowrins and funnelins, substrate hydrolysis follows a common general base/acid mechanism. A metal-bound solvent molecule ultimately performs the attack on the scissile peptide bond with the assistance of a strictly conserved glutamate residue.

Keywords:

• peptide bond hydrolysis
• three-dimensional X-ray crystal structure
• zinc enzyme
• catalysis

ACKNOWLEDGEMENTS

This study was supported by the following grants: BIO2006-02668, PSE-010000-2007-1, and CONSOLIDER-INGENIO 2010 Project “La Factoría de Cristalización” (CSD2006-00015) from Spanish ministries; EU FP6 Strep Project LSHG-2006-018830 “CAMP”; and EU FP7 Collaborative Project 223101 AntiPathoGN and 2005SGR00280 from the National Catalan Government. Robin Rycroft is thanked for helpful contributions to the manuscript and Frédéric Weber for assessment in conchology (http://pagesperso-orange.fr/frederic.weber/coquillages%20accueuil.htm).