Batch, Fed Batch Production and Characterization of Glutaminase Free L-Asparaginase II of Pectobacterium carotovorum MTCC 1428 in Escherichia coli - Advances in Microbiology

AiM > Vol.4 No.10, October 2014

Volume 4, Issue 10 (October 2014)

ISSN Print: 2165-3402 ISSN Online: 2165-3410

Google-based Impact Factor: 1.35 Citations

Batch, Fed Batch Production and Characterization of Glutaminase Free L-Asparaginase II of Pectobacterium carotovorum MTCC 1428 in Escherichia coli ()

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DOI: 10.4236/aim.2014.410072 4,910 Downloads 6,448 Views Citations

Author(s)

Rachna Goswami, Krishnamoorthy Hegde, Venkata Dasu Veeranki

Affiliation(s)

Biochemical Engineering Laboratory, Department of Biotechnology, Indian Institute of Technology Guwahati, Guwahati, Assam, India.

ABSTRACT

The present study describes the production optimization of recombinant L-asparaginase II of Pectobacterium carotovorum MTCC 1428 in Escherichia coli BL21 (DE3) at batch and fed batch bioreactor level. Production of recombinant L-asparaginase II in batch and fed batch mode was found to be 1.34 and 5.38 folds higher, respectively as compared to shake flask culture. SDS-PAGE and native PAGE of the purified enzyme revealed that molecular mass of the subunits and native enzyme are ~37.5 kDa and ~150 kDa, respectively. Optimum range of pH and temperature for hydrolysis of L-asparagine were found to be 7.5 - 8.5 and 47^°C - 52^°C, respectively. The recombinant enzyme is very specific for its natural substrate, L-asparagine. The activity of recombinant L-asparaginase II is improved by mono cations and diverse effectors including Na⁺, K⁺, L-cystine, L-histidine, glutathione and 2-mercaptoethanol whereas, it is moderately inhibited by different divalent cations and thiol group blocking reagent. The kinetic parameters K_m, V_max, k_cat and K_m/K_cat of purified recombinant L-asparaginase II were determined. The purified L-asparaginase II possesses no partial glutaminase activity, which is prerequisite to reduce the possibility of side effects during the course of anti-cancer therapy.

KEYWORDS

Batch Fermentation, Fed-Batch Fermentation, Recombinant L-Asparaginase II, Taguchi’s Method

Share and Cite:

Goswami, R. , Hegde, K. and Veeranki, V. (2014) Batch, Fed Batch Production and Characterization of Glutaminase Free L-Asparaginase II of Pectobacterium carotovorum MTCC 1428 in Escherichia coli. Advances in Microbiology, 4, 667-680. doi: 10.4236/aim.2014.410072.

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