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Purification and characterization of CHpro1, a thermotolerant, alkali-stable and oxidation-resisting protease of Chumathang hotspring

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  • Life & Medical Sciences
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Science Bulletin

Abstract

Metagenomic approaches are recently used for searching novel open reading frames (ORFs) coding enzymes employed in pharmaceutical, food industries, etc. In this study, a metagenomic library was constructed from Chumathang hotspring sediment DNA. The library consisted of approximately 9,000 clones and was screened for protease activity. A clone exhibiting protease activity was identified and named CHpro1. Sequencing of CHpro1 revealed that the ORF encoded a functional protein of 363 amino acids belonging to peptidase S8–S53 superfamily. CHpro1 shared 41 % sequence similarity with a reported protease (subtilase family) and 35 % structural similarity with the crystal structure of Pro-Tk sps. of Thermococcus kodarkaenasis. In silico modeling the 3D structure of CHpro1 showed that it has two beta sheets, 10 alpha helices and 11 strands. Catalytic triad prediction implied CHpro1 to be a serine protease. The optimum temperature and pH of the purified protease were found to be 80 °C and 11.0, respectively. The enzyme was active at 5 % concentration of hydrogen peroxide and retained 60 % of activity at 10 % concentration. The thermotolerant, alkalophilic and oxidant stable properties of the protease make it a potential candidate for biotechnological applications.

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Acknowledgments

This work was supported by Council of Scientific and Industrial Research (CSIR), New Delhi, Government of India (37(1545)/12/EMR-II) entitled “Exploring Microbial Diversity and Mining Novel Hydrolases from Brackish Water Lakes of Ladakh Region by Metagenomic Approach.” Reena Singh is thankful to Council of Scientific and Industrial Research (CSIR), Government of India, for the award of Senior Research fellowship.

Conflict of interest

The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. School of Biotechnology, Shri Mata Vaishno Devi University, Katra, 182320, India

    Reena Singh, Vishnu Kumar Gupta, Vijeshwar Verma & Shafaq Rasool

  2. Lovely Professional University, Phagwara, 144411, India

    Chirag Chopra

  3. Indian Institute of Toxicology Research, Lucknow, 223001, India

    Bashir Akhlaq

Authors
  1. Reena Singh
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  2. Chirag Chopra
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  3. Vishnu Kumar Gupta
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  4. Bashir Akhlaq
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  5. Vijeshwar Verma
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  6. Shafaq Rasool
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Corresponding author

Correspondence to Shafaq Rasool.

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Singh, R., Chopra, C., Gupta, V.K. et al. Purification and characterization of CHpro1, a thermotolerant, alkali-stable and oxidation-resisting protease of Chumathang hotspring. Sci. Bull. 60, 1252–1260 (2015). https://doi.org/10.1007/s11434-015-0834-8

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  • DOI: https://doi.org/10.1007/s11434-015-0834-8

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