|
KMID : 0613820060160071148
|
|
Journal of Life Science
2006 Volume.16 No. 7 p.1148 ~ p.1157
|
|
|
Cloning and Identification of Essential Residues for Thermostable β-Glucosidase (BglB) from Thermotoga maritima
|
|
Hong Su-Young
Cho Kye-Man
Kim Yong-Hee
Hong Sun-Joo
Cho Soo-Jeong
Cho Yong-Un
Kim Hoon
Yoon Han-Dae
|
|
|
Abstract
|
|
|
|
A hyperthermophilic bacterium Thermotoga maritima produced thermostable β-glucosidase. The gene encoding β-glucosidase from T. maritima MSB8 was cloned and expressed in Escherichia coli. The enzyme (BglB) hydrolyzed β-glucosidic linkages between glucose and alkyl, aryl of saccharide groups such as salicin, arbutin, and ρNPG. The insert DNA contained ORF with 2,166 bp encodes a 721 amino acids (calculated molecular mass of 80,964 and pI of 4.93). The amino acid sequence of BglB showed the similarity to family 3 glycosyl hydrolases. The molecular weight of the enzyme was estimated to be approximately 81 kDa by MUG-nondenaturing PAGE (4-methylumbelliferyl β-D-glucoside-nondenaturing polyacrylamide gel electophoresis) and SDS-PAGE. The β-glucosidase exhibited maximal activity at pH 7.0 and 80°C. By exchanging two possible residues (Glu-232 and Asp-242) to Ala by site-directed mutagenesis method, it was found that these were essential for enzymatic activity.
|
|
|
KEYWORD
|
|
|
Thermotoga maritima, β-glucosidase, essential residue, site-directed mutagenesis
|
|
|
FullTexts / Linksout information
|
|
|
|
|
|
Listed journal information
|
|
|