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Protein Engineering of a Metalloprotease in Order to Improve Organic Solvents Stability and Activity

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Abstract

Recently, improve the protease activity in the presence of organic solvents has been appreciated for the researchers. In the current study, we have tried to increase the organic solvent stability of salinovibrio proteolyticus protease (SVP) by site-directed mutagenesis. Five variants were constructed to substitute the surface charged, and polar amino acid residues in SVP with hydrophobic ones (T21V, Y23V, K30P, D25P and N248G) to examine the outcome of surface hydrophobicity on the enzyme efficiency in the presence of organic solvent. The catalytic efficiency of Y23V and N248G mutants not only increased about 1.8 and 2.6 folds in DMF and methanol but also increased it about 3.8 and 5.0 folds in isopropanol and n-propanol, compared to SVP. ∆∆G‡ values of Y23V and N248G variants, increased about 6.5 and 9.5 kcal mol−1 in DMF and methanol, and it improved about 13.6 and 16.6 kcal mol−1 in isopropanol and n-propanol, respectively. These results show that irreversible thermoinactivation rate of protease has a straight relationship with hydrophobicity of organic solvents.

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Authors and Affiliations

  1. Department of Biology, Faculty of Sciences, Shahid Bahonar University of Kerman, Kerman, Iran

    Arastoo Badoei-dalfard & Zahra Karami

  2. Department of Biochemistry, Faculty of Biological Science, Tarbiat Modares University, Tehran, Iran

    Khosro Khajeh

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  1. Arastoo Badoei-dalfard
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  2. Khosro Khajeh
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Correspondence to Arastoo Badoei-dalfard.

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Badoei-dalfard, A., Khajeh, K. & Karami, Z. Protein Engineering of a Metalloprotease in Order to Improve Organic Solvents Stability and Activity. Catal Lett 150, 1219–1229 (2020). https://doi.org/10.1007/s10562-019-03044-7

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