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Purification and characterization of an extracellular haloalkaline serine protease from the moderately halophilic bacterium, Bacillus iranensis (X5B)

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Abstract

This study reports the purification and characterization of an extracellular haloalkaline serine protease from the moderately halophilic bacterium, Bacillus iranensis, strain X5B. The enzyme was purified to homogeneity by acetone precipitation, ultrafiltration and carboxymethyl (CM) cation exchange chromatography, respectively. The purified protease was a monomeric enzyme with a relative molecular mass of 48–50 kDa and it was inhibited by PMSF indicating that it is a serine-protease. The optimum pH, temperature and NaCl concentration were 9.5, 35 °C and 0.98 M, respectively. The enzyme showed a significant tolerance to salt and alkaline pH. It retained approximately 50 % of activity at 2.5 M NaCl and about 70 % of activity at highly alkaline pH of 11.0; therefore, it was a moderately halophilic and also can be activated by metals, especially by Ca2+. The specific activity of the purified protease was measured to be 425.23 μmol of tyrosine/min per mg of protein using casein as a substrate. The apparent K m and V max values were 0.126 mM and 0.523 mM/min, respectively and the accurate value of k cat was obtained as 3.284 × 10−2 s−1. These special and important characteristics make this serine protease as valuable tool for industrial applications.

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Acknowledgments

We thank Sajjad Khani (Max-Planck-Institute for Metabolism Research) for the discussion and revision of the English in the manuscript. We also gratefully acknowledge the University of Guilan for their financial support.

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Authors and Affiliations

  1. Department of Biology, University of Guilan, Rasht, Iran

    Hossein Ghafoori & Mansoure Askari

  2. Molecular bank, Iranian Biological Resource Center (IBRC), ACECR, Tehran, Iran

    Sajjad Sarikhan

Authors
  1. Hossein Ghafoori
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  2. Mansoure Askari
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  3. Sajjad Sarikhan
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Correspondence to Hossein Ghafoori.

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Communicated by F. Robb.

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Ghafoori, H., Askari, M. & Sarikhan, S. Purification and characterization of an extracellular haloalkaline serine protease from the moderately halophilic bacterium, Bacillus iranensis (X5B). Extremophiles 20, 115–123 (2016). https://doi.org/10.1007/s00792-015-0804-8

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  • DOI: https://doi.org/10.1007/s00792-015-0804-8

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