Replacement of tyrosine 181 by phenylalanine in gentisate 1,2-dioxygenase I from Pseudomonas alcaligenes NCIMB 9867 enhances catalytic activities.
xlnE, encoding gentisate 1,2-dioxygenase (EC 1.13.11.4), from Pseudomonas alcaligenes (P25X) was mutagenized by site-directed mutagenesis. The mutant enzyme, Y181F, demonstrated 4-, 3-, 6-, and 16-fold increases in relative activity towards gentisate and 3-fluoro-, 4-methyl-, and 3-methylgentisate, respectively. The specific mutation conferred a 13-fold higher catalytic efficiency (k(cat)/Km) on Y181F towards 3-methylgentisate than that of the wild-type enzyme.
Tan
Chew Ling
CL
Programme in Environmental Microbiology, Department of Microbiology, Yong Loo Lin School of Medicine, National University of Singapore, 5 Science Drive 2, Singapore 117597.
Yeo
Chew Chieng
CC
Khoo
Hoon Eng
HE
Poh
Chit Laa
CL
D019943:Amino Acid Substitution; D002384:Catalysis; D049308:Dioxygenases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D005841:Gentisates / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D020125:Mutation, Missense; D044202:Pseudomonas alcaligenes / Q000201:enzymology* / Q000235:genetics; D013379:Substrate Specificity
D005841:Gentisates; D049308:Dioxygenases; C026247:gentisate 1,2-dioxygenase