Novel substrate specificity of a membrane-bound beta-glycosidase from the hyperthermophilic archaeon Pyrococcus horikoshii.
A beta-glycosidase gene homolog of Pyrococcus horikoshii (BGPh) was successfully expressed in Escherichia coli. The enzyme was localized in a membrane fraction and solubilized with 2.5% Triton X-100 at 85 degrees C for 15 min. The optimum pH was 6.0 and the optimum temperature was over 100 degrees C, respectively. BGPh stability was dependent on the presence of Triton X-100, the enzyme's half-life at 90 degrees C (pH 6.0) was 15 h. BGPh has a novel substrate specificity with k(cat)/K(m) values high enough for hydrolysis of beta-D-Glcp derivatives with long alkyl chain at the reducing end and low enough for the hydrolysis of beta-linked glucose dimer more hydrophilic than aryl- or alkyl-beta-D-Glcp.
Matsui
I
I
National Institute of Bioscience and Human-Technology, Tsukuba, Ibaraki, Japan. ikmatsui@nibh.go.jp
Sakai
Y
Y
Matsui
E
E
Kikuchi
H
H
Kawarabayasi
Y
Y
Honda
K
K
D000595:Amino Acid Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005962:Glucosylceramidase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008565:Membrane Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D019711:Pyrococcus / Q000201:enzymology*; D016415:Sequence Alignment; D013379:Substrate Specificity; D013696:Temperature
D008565:Membrane Proteins; D005962:Glucosylceramidase