Preparation, characterization, immobilization, and molecular docking analysis of a novel detergent-stable subtilisin-like serine protease from Streptomyces mutabilis strain TN-X30.

Autor: Mechri S; Laboratory of Microbial Biotechnology and Enzyme Engineering (LMBEE), Centre of Biotechnology of Sfax (CBS), University of Sfax, Road of Sidi Mansour Km 6, P.O. Box 1177, Sfax 3018, Tunisia., Allala F; Laboratory of Cellular and Molecular Biology (LCMB), Microbiology Team, Faculty of Biological Sciences (FSB), University of Sciences and Technology Houari Boumediene (USTHB), P.O. Box: 32, El Alia, Bab Ezzouar, 16111, Algiers, Algeria., Bouacem K; Laboratory of Cellular and Molecular Biology (LCMB), Microbiology Team, Faculty of Biological Sciences (FSB), University of Sciences and Technology Houari Boumediene (USTHB), P.O. Box: 32, El Alia, Bab Ezzouar, 16111, Algiers, Algeria., Hasnaoui I; Laboratory of Bioresources, Biotechnology, Ethnopharmacology and Health (LBBEH), Faculty of Sciences, Mohammed Premier University, Oujda 60000, Morocco; Univ Lyon, Université Lyon 1, Institut de Chimie et de Biochimie Moléculaires et Supramoléculaires (ICBMS), UMR 5246 CNRS, Génie Enzymatique, Membranes Biomimétiques et Assemblages Supramoléculaires (GEMBAS), Bât Raulin, 43 Bd du 11 Novembre 1918, F-69622 Villeurbanne Cedex, France., Gwaithan H; Department of Bioprocess Technology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia (UPM), Serdang, Kuala Lumpur, Malaysia., Chalbi TB; Laboratory of Microbial Biotechnology and Enzyme Engineering (LMBEE), Centre of Biotechnology of Sfax (CBS), University of Sfax, Road of Sidi Mansour Km 6, P.O. Box 1177, Sfax 3018, Tunisia., Saalaoui E; Laboratory of Bioresources, Biotechnology, Ethnopharmacology and Health (LBBEH), Faculty of Sciences, Mohammed Premier University, Oujda 60000, Morocco., Asehraou A; Laboratory of Bioresources, Biotechnology, Ethnopharmacology and Health (LBBEH), Faculty of Sciences, Mohammed Premier University, Oujda 60000, Morocco., Noiriel A; Univ Lyon, Université Lyon 1, Institut de Chimie et de Biochimie Moléculaires et Supramoléculaires (ICBMS), UMR 5246 CNRS, Génie Enzymatique, Membranes Biomimétiques et Assemblages Supramoléculaires (GEMBAS), Bât Raulin, 43 Bd du 11 Novembre 1918, F-69622 Villeurbanne Cedex, France., Abousalham A; Univ Lyon, Université Lyon 1, Institut de Chimie et de Biochimie Moléculaires et Supramoléculaires (ICBMS), UMR 5246 CNRS, Génie Enzymatique, Membranes Biomimétiques et Assemblages Supramoléculaires (GEMBAS), Bât Raulin, 43 Bd du 11 Novembre 1918, F-69622 Villeurbanne Cedex, France., Hacene H; Laboratory of Cellular and Molecular Biology (LCMB), Microbiology Team, Faculty of Biological Sciences (FSB), University of Sciences and Technology Houari Boumediene (USTHB), P.O. Box: 32, El Alia, Bab Ezzouar, 16111, Algiers, Algeria., Bouanane-Darenfed A; Laboratory of Cellular and Molecular Biology (LCMB), Microbiology Team, Faculty of Biological Sciences (FSB), University of Sciences and Technology Houari Boumediene (USTHB), P.O. Box: 32, El Alia, Bab Ezzouar, 16111, Algiers, Algeria., Le Roes-Hill M; Applied Microbial and Health Biotechnology Institute (AMHBI), Cape Peninsula University of Technology, P.O. Box 1906, Bellville 7535, South Africa., Jaouadi B; Laboratory of Microbial Biotechnology and Enzyme Engineering (LMBEE), Centre of Biotechnology of Sfax (CBS), University of Sfax, Road of Sidi Mansour Km 6, P.O. Box 1177, Sfax 3018, Tunisia. Electronic address: bassem.jaouadi@cbs.rnrt.tn.
Jazyk: angličtina
Zdroj: International journal of biological macromolecules [Int J Biol Macromol] 2022 Dec 01; Vol. 222 (Pt A), pp. 1326-1342. Date of Electronic Publication: 2022 Sep 20.
DOI: 10.1016/j.ijbiomac.2022.09.161
Abstrakt: We recently described the production of a detergent-biocompatible crude protease from Streptomyces mutabilis strain TN-X30. Here, we describe the purification, characterization, and immobilization of the serine alkaline protease (named SPSM), as well as the cloning, sequencing, and over-expression of its corresponding gene (spSM). Pure enzyme was obtained after ammonium sulphate precipitation followed by heat-treatment and Sephacryl® S-200 column purification. The sequence of the first 26 NH 2 -terminal residues of SPSM showed a high sequence identity to subtilisin-like serine proteases produced by actinobacteria. The spSM gene was heterologously expressed in Escherichia coli BL21(DE3)pLysS and E. coli BL21-AI™ strains using pTrc99A (rSPSM) and Gateway™ pDEST™ 17 [(His) 6 -tagged SPSM] vectors, respectively. Results obtained indicated that the (His) 6 -tagged SPSM showed the highest stability. The SPSM was immobilized using encapsulation and adsorption-encapsulation approaches and three different carriers. Features of SPSM in soluble and immobilized forms were analyzed by Fourier transform infrared (FTIR) spectroscopy in attenuated total reflection (ATR) mode, X-ray diffraction (XRD), zeta potential measurements, and field emission scanning electron microscopy (FE-SEM). The white clay and kaolin used in this study are eco-friendly binders to alginate-SPSM and show great potential for application of the immobilized SPSM in various industries. Molecular modeling and docking of N-succinyl-l-Phe-l-Ala-l-Ala-l-Phe-p-nitroanilide in the active site of SPSM revealed the involvement of 21 amino acids in substrate binding.
Competing Interests: Declaration of competing interest There are no conflicts of interest to declare.
(Copyright © 2022 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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