The method of integrated kinetics and its applicability to the exo-glycosidase-catalyzed hydrolyses of p-nitrophenyl glycosides

Anna S. Borisova, Sumitha K. Reddy, Dina R. Ivanen, Kirill S. Bobrov, Elena V. Eneyskaya, Georgy N. Rychkov, Mats Sandgren, Henrik Stålbrand, Michael L. Sinnott, Anna A. Kulminskaya, Konstantin A. Shabalin

Research output: Contribution to journalArticleScientificpeer-review

3 Citations (Scopus)

Abstract

In the present work we suggest an efficient method, using the whole time course of the reaction, whereby parameters kcat, Km and product KI for the hydrolysis of a p-nitrophenyl glycoside by an exo-acting glycoside hydrolase can be estimated in a single experiment. Its applicability was demonstrated for three retaining exo-glycoside hydrolases, β-xylosidase from Aspergillus awamori, β-galactosidase from Penicillium sp. and α-galactosidase from Thermotoga maritima (TmGalA). During the analysis of the reaction course catalyzed by the TmGalA enzyme we had observed that a non-enzymatic process, mutarotation of the liberated α-d-galactose, affected the reaction significantly.

Original languageEnglish
Pages (from-to)43-49
Number of pages7
JournalCarbohydrate Research
Volume412
DOIs
Publication statusPublished - 14 Aug 2015
MoE publication typeA1 Journal article-refereed

Keywords

  • Integrated kinetics
  • Mutarotation
  • Retaining glycoside hydrolase

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