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Prolonged Production and Aggregation Complexity of Cold-Active Lipase from Pseudomonas proteolytica (GBPI_Hb61) Isolated from Cold Desert Himalaya

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Abstract

Pseudomonas, being the common inhabitant of colder environments, are suitable for the production of cold-active enzymes. In the present study, a newly isolated strain of Pseudomonas from cold desert site in Indian Himalayan Region, was investigated for the production of cold-active lipase. The bacteria were identified as Pseudomonas proteolytica by 16S rDNA sequencing. Lipase production by bacteria was confirmed by qualitative assay using tributyrin and rhodamine-B agar plate method. The bacterium produced maximum lipase at 25 °C followed by production at 15 °C while utilizing olive, corn, as well as soybean oil as substrate in lipase production broth. Enzyme produced by bacteria was partially purified using ammonium sulphate fractionation. GBPI_Hb61 showed aggregation behaviour which was confirmed using several techniques including gel filtration chromatography, dynamic light scattering, and native PAGE. Molecular weight determined by SDS-PAGE followed by in-gel activity suggested two lipases of nearly similar molecular weight of ~50 kDa. The enzyme showed stability in wide range of pH from 5 to 11 and temperature up to 50 °C. The enzyme from GBPI_Hb61 exhibited maximum activity toward p-nitrophenyldecanoate (C10). The stability of enzyme was not affected with methanol while it retained more than 75% activity when incubated with ethanol, acetone, and hexane. The bacterium is likely to be a potential source for production of cold-active lipase with efficient applicability under multiple conditions.

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Abbreviations

pNP:

p-Nitrophenol

CHAPS:

3-[(3-Cholamidopropyl)dimethylammonio]-1-propane sulfonate

NP-40:

Nonylphenoxy polyethoxyethanol

Brij-35:

Polyethylene glycol dodecyl ether

SDS:

Sodium dodecyl sulphate

PEG:

Poly ethylene glycol

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Acknowledgements

The authors are thankful to Dr. P.P. Dhyani (Director, G.B. Pant National Institute of Himalayan Environment and Sustainable Development, India) for extending the facilities. Financial support from Ministry of Environment, Forest and Climate Change is gratefully acknowledged.

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Authors and Affiliations

  1. Biotechnological Applications, G B Pant National Institute of Himalayan Environment and Sustainable Development, Kosi-Katarmal, Almora, Uttarakhand, 263 643, India

    Rahul Jain & Anita Pandey

  2. Department of Microbiology, CSIR-Central Drug Research Institute, Lucknow, Uttar Pradesh, 226 031, India

    Mukesh Pasupuleti

  3. Department of Biotechnology, Kumaun University, Bhimtal Campus, Bhimtal, Uttarakhand, 263 136, India

    Veena Pande

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  1. Rahul Jain
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Jain, R., Pandey, A., Pasupuleti, M. et al. Prolonged Production and Aggregation Complexity of Cold-Active Lipase from Pseudomonas proteolytica (GBPI_Hb61) Isolated from Cold Desert Himalaya. Mol Biotechnol 59, 34–45 (2017). https://doi.org/10.1007/s12033-016-9989-z

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