Cloning, expression and characterization of L-asparaginase from Pseudomonas fluorescens for large scale production in E. coli BL21.

Vijay, Kishore and Nishita, K. P. and Manonmani, H. K. (2015) Cloning, expression and characterization of L-asparaginase from Pseudomonas fluorescens for large scale production in E. coli BL21. 3 Biotech, 5 (6). pp. 975-981.

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Abstract

L-Asparaginase (E.C. 3.5.1.1) is used as an antineoplastic drug in the treatment of acute lymphoblastic leukemia. L-Asparaginase from Pseudomonas fluorescens was cloned and overexpressed in E. coli BL21. The Enzyme was found to be a Fusion protein-asparaginase complex which was given a lysozyme treatment and sonication, and then was purified in a Sepharose 6B column. The enzymatic properties of the recombinant enzyme were studied and the kinetic parameters were determined with kilometre of 109.99 mM and Vmax of 2.88 lM/min. Recombinant enzyme showed pH optima at 6.3 and temperature optima at 34 �C. Asp gene was successfully cloned into E. coli BL21 which produced high level of asparaginase intracellularly with 85.25 % recovery of enzyme with a specific activity of 0.94 IU/mg protein. The enzyme was a tetramer with molecular weight of approximately 141 kDa.

Item Type:	Article
Uncontrolled Keywords:	L-Asparaginase Cloning E. coli Enzyme activity
Subjects:	600 Technology > 05 Chemical engineering > 03 Enzyme Biotechnology and Engineering
Divisions:	Fermentation Technology and Bioengineering
Depositing User:	Food Sci. & Technol. Information Services
Date Deposited:	08 Jan 2016 10:33
Last Modified:	08 Jan 2016 10:33
URI:	http://ir.cftri.res.in/id/eprint/12063

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