Glutamic acid 160 is the acid‐base catalyst of β‐xylosidase from Bacillus stearothermophilus T‐6: a family 39 glycoside hydrolase

FEBS Letters

Shoham, Gil²  Shoham, Yuval¹  Shulami, Smadar¹  Baasov, Timor³  Belakhov, Valery³  Bravman, Tsafrir¹  Mechaly, Adva¹ 

[1] Department of Food Engineering and Biotechnology, Technion Israel Institute of Technology, Haifa 32000, Israel;Department of Inorganic Chemistry and The Laboratory for Structural Chemistry and Biology, The Hebrew University of Jerusalem, Jerusalem 91904, Israel;Department of Chemistry, Technion Israel Institute of Technology, Haifa 32000, Israel

关键词: β-Xylosidase;    Glycoside hydrolase family 39;    Acid-base catalyst;    Mechanism;    Bacillus stearothermophilus;    PNPX;    p-nitrophenyl β-D-xylopyranoside;    ONPX;    o-nitrophenyl β-D-xylopyranoside;    2;    5-DNPX;    2′;    5′-dinitrophenyl β-D-xylopyranoside;    3;    4-DNPX;    3′;    4′-dinitrophenyl β-D-xylopyranoside;    SDS–PAGE;    sodium dodecyl sulfate–polyacrylamide gel electrophoresis;   

DOI  :  10.1016/S0014-5793(01)02371-7

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

A β-xylosidase from Bacillus stearothermophilus T-6 was cloned, overexpressed in Escherichia coli and purified to homogeneity. Based on sequence alignment, the enzyme belongs to family 39 glycoside hydrolases, which itself forms part of the wider GH-A clan. The conserved Glu160 was proposed as the acid-base catalyst. An E160A mutant was constructed and subjected to steady state and pre-steady state kinetic analysis together with azide rescue and pH activity profiles. The observed results support the assignment of Glu160 as the acid-base catalytic residue.

【 授权许可】

Unknown