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Enhanced secretory production of hemolysin-mediated cyclodextrin glucanotransferase in escherichia coli by random mutagenesis of the abc transporter system

Md. Illias , Rosli and Kheng Oon, Low and Mahadi, Nor Muhammad and Abdul Rahim, Raha and Rabu, Amir and Abu Bakar, Farah Diba and Abdul Murad, Abdul Munir (2010) Enhanced secretory production of hemolysin-mediated cyclodextrin glucanotransferase in escherichia coli by random mutagenesis of the abc transporter system. Journal of Biotechnology, 150 (4). 453 - 459. ISSN 0168-1656

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Official URL: http://dx.doi.org/10.1016/j.jbiotec.2010.10.001

Abstract

The hemolysin transport system was found to mediate the release of cyclodextringlucanotransferase (CGTase) into the extracellular medium when it was fused to the C-terminal 61 amino acids of HlyA (HlyAs61). To produce an improved-secretion variant, the hly components (hlyAs, hlyB and hlyD) were engineered by directed evolution using error-prone PCR. Hly mutants were screened on solid LB-starch plate for halo zone larger than the parent strain. Through screening of about 1 × 104Escherichiacoli BL21(DE3) transformants, we succeeded in isolating five mutants that showed a 35–217% increase in the secretion level of CGTase-HlyAs61 relative to the wild-type strain. The mutation sites of each mutant were located at HlyB, primarily along the transmembrane domain, implying that the corresponding region was important for the improved secretion of the target protein. In this study we describe the finding of novel site(s) of HlyB responsible for enhancing secretion of CGTase in E. coli.

Item Type:Article
Uncontrolled Keywords:alpha-hemolysin transport system, cyclodextringlucanotransferase, directed evolution, extracellular secretion, escherichiacoli
Subjects:Q Science > QD Chemistry
Divisions:Chemical Engineering
ID Code:26167
Deposited By: Liza Porijo
Deposited On:28 Jun 2012 00:40
Last Modified:06 Jul 2012 04:13

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