title: Mutation of a conserved tryptophan in the chitin-binding cleft of Serratia marcescens chitinase A enhances transglycosylation.
abstract: Family 18 chitinases have the signature peptide DGXDXDXE forming the fourth beta-strand in the (beta/alpha)8-barrel of their catalytic domain. The carboxyl-end glutamic acid, E315 in Serratia marcescens chitinase A, serves as the acid/base during chitin hydrolysis, and the side-chain of the preceding aspartic acid, D313, helps to position correctly the N-acetyl moiety of the glycosyl sugar undergoing hydrolysis. Chitin substrates are bound within a long cleft across the top of the barrel, whose floor consists of aromatic residues that hydrophobically stack with every other GlcNAc. Alanine substitution of the conserved Trp167 at the -3 subsite in Serratia marcescens chitinase A enhanced transglycosylation. Higher oligosaccharides were formed from both chitin tetra- and pentasaccharide, and the only hydrolytic product from chitin trisaccharide was the disaccharide. Greater retention of the glycosyl fragment at the active site of the -3 mutant of Serratia marcescens chitinase A might favor transglycosylation due to a stabilized conformation of its D313.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of South Alabama, AL 36688, USA. naronson@jaguar1.usouthal.edu
authors.forename: Nathan N
authors.initials: NN
authors.lastname: Aronson
authors.forename: Brian A
authors.initials: BA
authors.lastname: Halloran
authors.forename: Mikhail F
authors.initials: MF
authors.lastname: Alexeyev
authors.forename: Xiaoyin E
authors.initials: XE
authors.lastname: Zhou
authors.forename: Yujun
authors.initials: Y
authors.lastname: Wang
authors.forename: Edward J
authors.initials: EJ
authors.lastname: Meehan
authors.forename: Liqing
authors.initials: L
authors.lastname: Chen
mesh_terms: D001665:Binding Sites; D002686:Chitin / Q000378:metabolism*; D002688:Chitinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017124:Conserved Sequence / Q000235:genetics*; D006031:Glycosylation; D006868:Hydrolysis; D007700:Kinetics; D015394:Molecular Structure; D009154:Mutation / Q000235:genetics*; D012706:Serratia marcescens / Q000201:enzymology* / Q000235:genetics; D013379:Substrate Specificity; D014364:Tryptophan / Q000235:genetics / Q000378:metabolism*
chemical_list: D002686:Chitin; D014364:Tryptophan; D002688:Chitinases