Paper
The New Mechanism of Substrate Recognition by Metallocarboxypeptidases
- Authors:
- Valery Kh. Akparov; Vladimir I. Timofeev; Ilyas G. Khaliullin; Vytas ?vedas; Galina G. Chestukhina; Inna P. Kuranova
- Abstract
- Carboxypeptidase T from Thermoactinomyces vulgaris (CpT) contains aspartic acid residue on the bottom of primary specificity pocket, like carboxypeptidase B (CpB), but CpT is able to chip off hydrophobic C-terminal amino acid residues like carboxypeptidase A (CpA) as well as positively-charged (like CpB) and even negatively-charged ones. To clarify the CpT substarte recognition mechanism, the 3-D structures of complexes of CpT with sulfamide based phenylalanine and arginine transition-state analogs were determined. It is shown that conservative Leu203, (CpA numbering) Leu247 (CpA numbering) is structural determinants of hydrophobic, and Asp256 of positively charged substrates. It was shown also that Asp253 is uncharged. Variant CpT Asp256Asn has CpA-like selectivity, and variant CpT Leu247Asn can’t chip off negative charged residues. The role of calcium ions in allosteric regulation of CpT selectivity (Akparov et al., Crystallography Reports. 56(4), 596–602 as well as a role of fixed water molecules in substrate binding were revealed also. Side-chain depended shift (0.1 Ǻ) of ligand relative to Zn atom induce the conformation rearrangements of Glu270 (0.4Ǻ) and Arg127 (0.1 Ǻ) in catalytic center of CpT and as a result S1’-subsite can regulate the catalysis efficiency. This is a possible new mechanism of substrate recognition in metallocarboxypeptidases.
- Keywords
- Metallocarboxypeptidases; Carboxypeptidase T; Thermoactinomyces Vulgaris
- StartPage
- 200
- EndPage
- 200
- Doi