title: Role of Tyr-435 of Vibrio harveyi chitinase A in chitin utilization.
abstract: Vibrio harveyi chitinase A or VhChiA (EC.3.2.1.14) is a member of GH-18 chitinases that catalyzes chitin degradation from marine biomaterials. Our earlier structural data of VhChiA suggested that Tyr-435 marks the ending of subsite +2 and may influence binding of the interacting substrate at the aglycone binding sites. This study reports the effects of Tyr-435 using site-directed mutagenesis technique. Mutation of Tyr-435 to Ala (mutant Y435A) enhanced both binding and catalytic efficiency of VhChiA, whereas substitution of Tyr-435 to Trp (mutant Y435W) lessened the ability of the enzyme to bind and hydrolyze chitin substrates. The increased activity of Y435A can be explained by partial removal of a steric clash around subsite (+2), thereby allowing a chitin chain to move beyond or to access the enzyme's active site from the aglycone side more straightforwardly.
authors.affiliation: Biochemistry-Electrochemistry Research Unit, Schools of Chemistry and Biochemistry, Institute of Science, Suranaree University of Technology, Nakhon Ratchasima, Thailand.
authors.forename: Natchanok
authors.initials: N
authors.lastname: Sritho
authors.forename: Wipa
authors.initials: W
authors.lastname: Suginta
mesh_terms: D019943:Amino Acid Substitution; D002686:Chitin / Q000378:metabolism*; D002688:Chitinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006868:Hydrolysis; D007700:Kinetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D011487:Protein Conformation; D013379:Substrate Specificity; D014443:Tyrosine; D014733:Vibrio / Q000201:enzymology*
chemical_list: D002686:Chitin; D014443:Tyrosine; D002688:Chitinases