Transglycosylation by barley α-amylase 1

János A. Mótyán, Erika Fazekas, Haruhide Mori, Birte Svensson, Péter Bagossi, Lili Kandra, Gyöngyi Gyémánt

Research output: Article

6 Citations (Scopus)

Abstract

The transglycosylation activity of barley α-amylase 1 (AMY1) and active site AMY1 subsite mutant enzymes was investigated. We report here the transferase ability of the V47A, V47F, V47D and S48Y single mutants and V47K/S48G and V47G/S48D double mutant AMY1 enzymes in which the replaced amino acids play important role in substrate binding at subsites at -3 through -5. Although mutation increases the transglycosylation activity of enzymes, in the presence of acceptors the difference between wild type and mutants is not so significant. Oligomer transfer reactions of AMY1 wild type and its mutants were studied using maltoheptaose and maltopentaose donors and different chromophore containing acceptors. The conditions for the chemoenzymatic synthesis of 4-methylumbelliferyl-α-d-maltooligosaccharides (MU-α-d-MOSs) were optimized using 4-methylumbelliferyl-β-d-glucoside as acceptor and maltoheptaose as donor. 4-Methylumbelliferyl-α-d-maltoside, -maltotrioside, -maltotetraoside and -maltopentaoside have been synthesized. Products were identified by MALDI-TOF MS. 1H and 13C NMR analyses showed that AMY1 V47F preserved the stereo-and regioselectivity. The produced MU-α-d-MOSs of degree of polymerization DP 2, DP 3 and DP 5 were successfully applied to detect activity of Bacillus stearothermophilus maltogenic α-amylase, human salivary α-amylase and Bacillus licheniformis α-amylase, respectively in a fast and simple fluorometric assay.

Original languageEnglish
Pages (from-to)229-237
Number of pages9
JournalJournal of Molecular Catalysis B: Enzymatic
Volume72
Issue number3-4
DOIs
Publication statusPublished - nov. 1 2011

ASJC Scopus subject areas

  • Catalysis
  • Bioengineering
  • Biochemistry
  • Process Chemistry and Technology

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