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Novel glutaminase free l-asparaginase from Nocardiopsis alba NIOT-VKMA08: production, optimization, functional and molecular characterization

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Abstract

Studies were carried out for the optimization and production of novel extracellular glutaminase-free l-asparaginase from Nocardiopsis alba NIOT-VKMA08. Among the tested carbon and nitrogen sources, maximum l-asparaginase production was observed with a combination of l-asparagine and maltose (1.5 %) and twofold increase in yield (18.47 IU mL−1) was observed with newly optimized NIOT-asparaginase medium. Activity of the purified enzyme was moderately inhibited by various divalent cations and thiol group blocking reagents, with K m and V max of 0.127 mM and 5.50 U µg−1. Optimum pH and temperature of purified l-asparaginase for the hydrolysis of l-asparagine was 8.0 and 37 °C, respectively. The enzyme inhibited polyacrylamide formation in 10 % solution and it was very specific for its natural substrate l-asparagine. Partial glutaminase activity was not detected, which could reduce the possibility of side effects during cancer therapy. l-Asparaginase biosynthesis gene (ansA) was cloned and transformed in E. coli JM109. The ansA gene sequence reported in this study contains several base substitutions with that of reported sequences in GenBank, resulting in altered amino acid sequences of the translated protein.

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Acknowledgments

The authors gratefully acknowledge the financial support by the Earth System Sciences Organization (ESSO), Ministry of Earth Sciences, Government of India, New Delhi to conduct this research. The authors are thankful to Dr. M. A. Atmanand, Director, ESSO-National Institute of Ocean Technology (ESSO-NIOT), Chennai for the constant support and encouragement  to carry out this research work. The authors are profoundly thankful to Prof. T. Subramoniam, D.Sc., F.N.A. and Dr. M. Vijayakumaran for their critical comments and suggestions to improve this manuscript, and Dr. Toms C. Joseph, Senior Scientist, Microbiology, Fermentation and Biotechnology Division, Central Institute of Fisheries Technology (CIFT), Kochi, India for DNA sequencing, in silico sequence analysis and molecular studies.

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Authors and Affiliations

  1. Andaman and Nicobar Centre for Ocean Science and Technology, Earth System Sciences Organization, National Institute of Ocean Technology (ESSO-NIOT), Dollygunj, Port Blair, 744 103, Andaman and Nicobar Islands, India

    Balakrishnan Meena, Lawrance Anburajan, Palaiya Sukumaran Dheenan, Mehmuna Begum & Nambali Valsalan Vinithkumar

  2. Marine Biotechnology Group, ESSO-NIOT, Ministry of Earth Sciences, Govt. of India, Chennai, 600 100, India

    Gopal Dharani & Ramalingam Kirubagaran

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  1. Balakrishnan Meena
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  2. Lawrance Anburajan
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  3. Palaiya Sukumaran Dheenan
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  4. Mehmuna Begum
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  5. Nambali Valsalan Vinithkumar
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  6. Gopal Dharani
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  7. Ramalingam Kirubagaran
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Correspondence to Balakrishnan Meena.

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B. Meena and L. Anburajan contributed equally to this work.

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Meena, B., Anburajan, L., Dheenan, P.S. et al. Novel glutaminase free l-asparaginase from Nocardiopsis alba NIOT-VKMA08: production, optimization, functional and molecular characterization. Bioprocess Biosyst Eng 38, 373–388 (2015). https://doi.org/10.1007/s00449-014-1277-3

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