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Cloning, extracellular expression and characterization of predominant β-CGTase from Bacillus sp. G1 in E. coli


Citation

Ong, Rui Min and Goh, Kian Mau and Mahadi, Nor Muhammad and Hassan, Osman and Raja Abdul Rahman, Raja Noor Zaliha and Md. Illias, Rosli (2008) Cloning, extracellular expression and characterization of predominant β-CGTase from Bacillus sp. G1 in E. coli. Journal of Industrial Microbiology & Biotechnology, 35 (12). pp. 1705-1714. ISSN 1367-5435; ESSN: 1476-5535

Abstract

The cyclodextrin glucanotransferase (CGTase, EC 2.4.1.19) gene from Bacillus sp. G1 was successfully isolated and cloned into Escherichia coli. Analysis of the nucleotide sequence revealed the presence of an open reading frame of 2,109 bp and encoded a 674 amino acid protein. Purified CGTase exhibited a molecular weight of 75 kDa and had optimum activity at pH 6 and 60 degrees C. Heterologous recombinant protein expression in E. coli is commonly problematic causing intracellular localization and formation of inactive inclusion bodies. This paper shows that the majority of CGTase was secreted into the medium due to the signal peptide of Bacillus sp. G1 that also works well in E. coli, leading to easier purification steps. When reacted with starch, CGTase G1 produced 90% β-cyclodextrin (CD) and 10% γ-CD. This enzyme also preferred the economical tapioca starch as a substrate, based on kinetics studies. Therefore, CGTase G1 could potentially serve as an industrial enzyme for the production of β-CD.


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Additional Metadata

Item Type: Article
Divisions: Faculty of Biotechnology and Biomolecular Sciences
DOI Number: https://doi.org/10.1007/s10295-008-0462-2
Publisher: Springer
Keywords: Bacillus sp. G1; Cyclodextrin; Cyclodextrin glucanotransferase; Extracellular expression; Predominant β-CGTase; Signal peptide
Depositing User: Nida Hidayati Ghazali
Date Deposited: 24 Jul 2013 01:55
Last Modified: 28 Sep 2016 04:38
Altmetrics: http://www.altmetric.com/details.php?domain=psasir.upm.edu.my&doi=10.1007/s10295-008-0462-2
URI: http://psasir.upm.edu.my/id/eprint/13611
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