Characterization of an extracellular thermophilic alkaline esterase produced by Bacillus subtilis DR8806 |
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| Institution: | 1. Cellular and Molecular Research Group, Institute of Biotechnology, Ferdowsi University of Mashhad, Mashhad, Iran;2. Department of Chemistry, Faculty of Science, Ferdowsi University of Mashhad, Mashhad, Iran;1. State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China;2. Institute of Life Sciences, Jiangsu University, Zhenjiang 212013, China;1. Laboratoire de Génie Chimique, CNRS—Université de Toulouse (INPT), 4 allée Emile Monso, BP 84234, 31432 Toulouse, France;2. Institut National de la Recherche Agronomique (INRA), Laboratoire de Biotechnologie de l''Environnement, Avenue des Etangs, 11100 Narbonne, France;1. Laboratory of Immunocytochemistry, Program of Anatomy and Developmental Biology, School of Medicine, University of Chile, Chile;2. Departamento de Biología, Universidad de Tarapacá, Chile;1. Jiangsu Key Laboratory of Crop Genetics and Physiology/Co-Innovation Center for Modern Production Technology of Grain Crops, Yangzhou University, Yangzhou, China;2. State Key Laboratory of Crop Genetics and Germplasm Enhancement, College of Horticulture, Nanjing Agricultural University, Weigang, Nanjing, China |
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| Abstract: | This work is a report of the characterization of an alkaline lipolytic enzyme isolated from Bacillus subtilis DR8806. The extracellular extract was concentrated using ammonium sulfate, and ultrafiltration. The active enzyme was purified by Q-sepharose ion exchange chromatography. The molecular mass of the enzyme was estimated to be 60.25 kDa based on SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis). The optimum pH and temperature of this enzyme were observed to be 8.0 and 50 °C, respectively. The enzyme exhibited a half-life of 72 min at its optimum temperature. It was stable in the presence of metal ions (10 mM) such as Ca2+, K+ and Na+, whereas Cu2+, Fe2+, Zn2+, Mn2+, Co2+, Mg2+ and Hg2+ were found to have inhibitory effects. However, the enzyme activity was not affected significantly by 1% Triton X-100. The study of substrate specificity showed that the purified enzyme has a preferential specificity for small ester of p-nitrophenyl acetate (C2), and it was the most efficiently hydrolyzed substrate as compared to the other esters. The kinetic parameters showed that the enzyme has Km of 4.2 mM and Vmax of 151 μmol min?1 mg?1 for p-nitrophenyl acetate. The hydrolysis rates of the fluorescence substrates were increased in the presence of the purified enzyme. Regarding the features of the enzyme, it may be utilized as a novel candidate for industrial applications. |
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