Saturation-transfer-difference NMR to characterize substrate binding recognition and catalysis of two broadly specific glycoside hydrolases

Lothar Brecker, Grit D. Straganz, Catrin E. Tyl, Walter Steiner, Bernd Nidetzky

Research output: Contribution to journalArticlepeer-review

15 Scopus citations

Abstract

Saturation-transfer-difference NMR spectroscopy (STD NMR) is used to delineate noncovalent enzyme-substrate interactions of β-glycosidases from Pyrococcus furiosus and Aspergillus fumigatus under binding-only conditions at low temperatures, and during catalysis. Glucopyranosyl and galactopyranosyl moieties display a distinct pattern of multiple contacts with each active site, revealing enzyme-specific elements of recognition and portraying the global binding effect caused by single-site modification of the substrate, at carbon 4. The glucopyranose leaving group of cellobiose or lactose shows small relative STD effects except for the anomeric carbon, particularly in the α-form. Its replacement in β-glucosides by an alcohol leaving group strongly affects sugar binding in the proximal enzyme subsite. A combination of STD effects of substrate and product, produced by the catalytic event or added exogenously, characterizes subsite binding during cellobiose hydrolysis.

Original languageEnglish (US)
Pages (from-to)85-89
Number of pages5
JournalJournal of Molecular Catalysis B: Enzymatic
Volume42
Issue number3-4
DOIs
StatePublished - Nov 2 2006

Bibliographical note

Copyright:
Copyright 2008 Elsevier B.V., All rights reserved.

Keywords

  • Enzyme-substrate interaction
  • Glycoside hydrolysis
  • Noncovalent binding recognition
  • STD NMR
  • β-Glycosidase

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