Stereochemistry of family 52 glycosyl hydrolases: A β-xylosidase from Bacillus stearothermophilus T-6 is a retaining enzyme

Tsafrir Bravman, Gennady Zolotnitsky, Smadar Shulami, Valery Belakhov, Dmitry Solomon, Timor Baasov, Gil Shoham, Yuval Shoham*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

42 Scopus citations

Abstract

A β-xylosidase from Bacillus stearothermophilus T-6 assigned to the uncharacterized glycosyl hydrolase family 52 was cloned, overexpressed in Escherichia coli and purified. The enzyme showed maximum activity at 65°C and pH 5.6-6.3. The stereochemistry of the hydrolysis of p-nitrophenyl β-D-xylopyranoside was followed by 1H-nuclear magnetic resonance. Time dependent spectrum analysis showed that the configuration of the anomeric carbon was retained, indicating that a retaining mechanism prevails in family 52 glycosyl hydrolases. Sequence alignment and site-directed mutagenesis enabled the identification of functionally important amino acid residues of which Glu337 and Glu413 are likely to be the two key catalytic residues involved in enzyme catalysis.

Original languageAmerican English
Pages (from-to)39-43
Number of pages5
JournalFEBS Letters
Volume495
Issue number1-2
DOIs
StatePublished - 20 Apr 2001

Bibliographical note

Funding Information:
This study was supported by Grants from the Israel Science Foundation (administered by the Israel Academy of Sciences and Humanities, Jerusalem) (no. 676/00 to G.S. and Y.S.), and from the United States–Israel Binational Science Foundation (BSF) (no. 96-178 to Y.S.), Jerusalem, Israel. Additional support was provided by the Fund for the Promotion of Research at the Technion, and by the Otto Meyerhof Center for Biotechnology, established by the Minerva Foundation (Munich, Germany). V.B. acknowledges the financial support by the Center of Absorption in Science, the Ministry of Immigration Absorption and the Ministry of Science and Arts, Israel (Kamea Program).

Keywords

  • Bacillus stearothermophilus
  • Catalytic residue
  • Glycosyl hydrolase family 52
  • Nuclear magnetic resonance
  • Retaining mechanism
  • β-Xylosidase

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