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Glucansucrase Gtf180-Delta N of Lactobacillus reuteri 180: enzyme and reaction engineering for improved glycosylation of non-carbohydrate molecules

Devlamynck, T., Te Poele, E. M., Meng, X., van Leeuwen, S. S. & Dijkhuizen, L., Sep-2016, In : Applied Microbiology and Biotechnology. 100, 17, p. 7529-7539 11 p.

Research output: Contribution to journalArticleAcademicpeer-review

Glucansucrases have a broad acceptor substrate specificity and receive increased attention as biocatalysts for the glycosylation of small non-carbohydrate molecules using sucrose as donor substrate. However, the main glucansucrase-catalyzed reaction results in synthesis of alpha-glucan polysaccharides from sucrose, and this strongly impedes the efficient glycosylation of non-carbohydrate molecules and complicates downstream processing of glucosylated products. This paper reports that suppressing alpha-glucan synthesis by mutational engineering of the Gtf180-Delta N enzyme of Lactobacillus reuteri 180 results in the construction of more efficient glycosylation biocatalysts. Gtf180-Delta N mutants (L938F, L981A, and N1029M) with an impaired alpha-glucan synthesis displayed a substantial increase in monoglycosylation yields for several phenolic and alcoholic compounds. Kinetic analysis revealed that these mutants possess a higher affinity for the model acceptor substrate catechol but a lower affinity for its mono-alpha-d-glucoside product, explaining the improved monoglycosylation yields. Analysis of the available high resolution 3D crystal structure of the Gtf180-Delta N protein provided a clear understanding of how mutagenesis of residues L938, L981, and N1029 impaired alpha-glucan synthesis, thus yielding mutants with an improved glycosylation potential.

Original languageEnglish
Pages (from-to)7529-7539
Number of pages11
JournalApplied Microbiology and Biotechnology
Volume100
Issue number17
Publication statusPublished - Sep-2016

    Keywords

  • Glycosylation, Glucansucrase, Catechol, Lactobacillus reuteri, Acceptor reaction, Enzyme engineering, SUCROSE PHOSPHORYLASE, ALPHA-GLUCOSIDE, ACTIVE-SITE, SPECIFICITY, GLUCOSYLTRANSFERASE, TRANSGLYCOSYLATION, RESVERATROL, ANALOGS, GTF180

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