The effects of the surface-exposed residues on the binding and hydrolytic activities of Vibrio carchariae chitinase A.

BMC Biochemistry
Supansa PantoomWipa Suginta

Abstract

Vibrio carchariae chitinase A (EC3.2.1.14) is a family-18 glycosyl hydrolase and comprises three distinct structural domains: i) the amino terminal chitin binding domain (ChBD); ii) the (alpha/beta)8 TIM barrel catalytic domain (CatD); and iii) the alpha + beta insertion domain. The predicted tertiary structure of V. carchariae chitinase A has located the residues Ser33 & Trp70 at the end of ChBD and Trp231 & Tyr245 at the exterior of the catalytic cleft. These residues are surface-exposed and presumably play an important role in chitin hydrolysis. Point mutations of the target residues of V. carchariae chitinase A were generated by site-directed mutagenesis. With respect to their binding activity towards crystalline alpha-chitin and colloidal chitin, chitin binding assays demonstrated a considerable decrease for mutants W70A and Y245W, and a notable increase for S33W and W231A. When the specific hydrolyzing activity was determined, mutant W231A displayed reduced hydrolytic activity, whilst Y245W showed enhanced activity. This suggested that an alteration in the hydrolytic activity was not correlated with a change in the ability of the enzyme to bind to chitin polymer. A mutation of Trp70 to Ala caused the most severe loss in b...Continue Reading

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Citations

Nov 13, 2009·Applied Microbiology and Biotechnology·Daniel GuillénRomina Rodríguez-Sanoja
Dec 24, 2011·Applied Biochemistry and Biotechnology·Natchanok Sritho, Wipa Suginta
Jul 2, 2009·Journal of Chemical Biology·Wipa SugintaHeino Prinz
Jun 25, 2010·Critical Reviews in Biotechnology·Chilukoti NeerajaAppa Rao Podile
Aug 16, 2014·Bioscience, Biotechnology, and Biochemistry·Paknisa SirimontreeTamo Fukamizo
Feb 18, 2017·Marine Drugs·Alexander J WinklerMarta Berrocal-Lobo
May 3, 2011·The Journal of Biological Chemistry·Supansa PantoomWipa Suginta
Dec 12, 2012·Bioscience, Biotechnology, and Biochemistry·Wipa Suginta, Natchanok Sritho

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Methods Mentioned

BETA
PCR

Software Mentioned

Superpose
TrEMBL
Genedoc
Model
PredicProtein
QuickChange
DNASTAR package
CCP4
DNASTAR
Pdb Viewer

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