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Crystal structure and identification of a key amino acid for glucose tolerance, substrate specificity, and transglycosylation activity of metagenomic β‐glucosidase Td2F2
Author(s) -
Matsuzawa Tomohiko,
Jo Toshinori,
Uchiyama Taku,
Manninen Jenny A.,
Arakawa Takatoshi,
Miyazaki Kentaro,
Fushinobu Shinya,
Yaoi Katsuro
Publication year - 2016
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.13743
Subject(s) - mutant , hydrolase , chemistry , biochemistry , mutagenesis , substrate (aquarium) , glycoside hydrolase , stereochemistry , enzyme , biology , ecology , gene
β-Glucosidase Td2F2 isolated from a compost metagenome has high glucose tolerance and transglycosylation activity. In this study, we determined the high-resolution crystal structure of Td2F2. It has a unique structure at the -1 subsite that is important for substrate specificity but not for glucose tolerance. To elucidate the mechanism(s) of glucose tolerance, we isolated a glucose-sensitive Td2F2 mutant using random mutagenesis. In this mutant, Asn223 residue located between subsites +1 and +2 was mutated. The Asn223 mutation resulted in reduced glucose tolerance and transglycosylation activity, and drastically changed substrate specificity. These results indicate that the structure between subsites +1 and +2 is critical for the glucose tolerance and substrate specificity of Td2F2. Our findings shed light on the glucose tolerance and transglycosylation activity mechanisms of glycoside hydrolase family 1 β-glucosidases.