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Genetic and biochemical characterization of thermophilic β-cyclodextrin glucanotransferase from Gracilibacillus alcaliphilus SK51.001.

Mohammed Abdalla,Hinawi A. M. Hassanin,Xiaolin Yao,Muhammad Waheed Iqbal,E. Karrar,B. Jiang

Published 2020 in The Journal of the Science of Food and Agriculture

ABSTRACT

BACKGROUND A strain, Gracilibacillus alcaliphilus SK51.001, which produces β-CGTase (β-cyclodextrin glucanotransferase) (EC 2.4.1.19), was screened and isolated from Sudanese soil. The objective of this study was to sequence and characterize the β-CGTase gene from G. alcaliphilus SK51.001. RESULTS According to 16S rRNA analysis and the morphological shape the strain was identified as Gracilibacillus alcaliphilus. The β-CGTase gene was successfully cloned, sequenced and expressed in Escherichia coli BL21. This gene showed 706 amino acid residues with 33 amino acids as a signal peptide. The active site residues of G. alcaliphilus SK51.001CGTase were described using enzyme modeling and docking with the products. The estimated molecular mass of G. alcaliphilus SK51.001CGTase was approximately 74 kDa as determined by SDS-PAGE, while evaluation of the gel filtration showed approximately 85 kDa, which means G. alcaliphilus SK51.001CGTase is a monomer. The optimum temperature and pH of G. alcaliphilus SK51.001CGTase were 60 °C and 7.0 respectively. G. alcaliphilus SK51.001CGTase was comparatively stable at pH between 6.0-9.0 and temperature 30-50 °C. The activity of G. alcaliphilus SK51.001CGTase was increased by Ni2+ , and Co2+ but inhibited by Al3+ and Fe3+ . The kinetic parameters of Km and Vmax were 2068.52 μg mL-1 and 0.13 μmol mL-1 min-1 , respectively. CONCLUSION G. alcaliphilus SK51.001CGTase could hydrolyze soluble starch into α-, β-, and γ-cyclodextrin in ratio of 2: 83: 15 % respectively, this high ratio production of β-CD could allow the enzyme to be used in β-CD production. This article is protected by copyright. All rights reserved.

PUBLICATION RECORD

  • Publication year

    2020

  • Venue

    The Journal of the Science of Food and Agriculture

  • Publication date

    2020-11-22

  • Fields of study

    Biology, Medicine, Chemistry, Environmental Science

  • Identifiers
    DOI 10.1002/jsfa.10960PMID 33222223
  • External record

    Open on Semantic Scholar

  • Source metadata

    Semantic Scholar, PubMed

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