Evaluation of plant lepidium peroxidase enzyme stability at different PH

سال انتشار: 1399
محل انتشار: بیست و یکمین کنگره ملی و نهمین کنگره بین المللی زیست شناسی ایران
کد COI اختصاصی: BIOCONF21_1029
زبان مقاله: انگلیسی
تعداد مشاهده: 143

نویسندگان

Department of Biotechnology, Institute of Science and High technology and Environmental Sciences, Graduate University of Advanced Technology, Kerman, Iran

Ali Riahi-Madvar

Department of Molecular and Cell Biology, Faculty of Basic Sciences, Kosar University of Bojnord, Bojnord, Iran

Mojtaba Mortazavi

Department of Biotechnology, Institute of Science and High technology and Environmental Sciences, Graduate University of Advanced Technology, Kerman, Iran

Safa Lotfi

Department of Biotechnology, Institute of Science and High technology and Environmental Sciences, Graduate University of Advanced Technology, Kerman, Iran

چکیده

Peroxidases (EC ۱.۱۱.۱.۷) are oxidoreductases that catalyze the reduction of peroxides such as hydrogen peroxide (H۲O۲) and the oxidation of a variety of organic and inorganic compounds. Most commercial applications of peroxides are in analytical diagnostics such as biosensors and immunoassay. In this study, the main purpose was to evaluate the stability of plant Lepidium draba peroxidase (LDP) at different pH ranges. In this study, after culturing bacteria containing pET۲۸a vector, which carries the plant peroxidase gene LDP, using IPTG inducer, expression was increased for ۴ hours at ۱۸ °C. Protein purification was performed using Ni-NTA affinity chromatography column, and using imidazole-containing buffer. Protein purity was determained by SDS-PAGE gel electrophoresis and its concentration was measured by Bradford method and its activity was measured in the presence of TMB and H۲O۲ substrates at ۶۵۳ nm using a spectrophotometer. Then enzyme activity was measured at different pH (۵-۹) with three repetitions. The results of gel electrophoresis showed that the protein was purified with high purity and has the highest activity in the pH ranges of ۶ to ۶/۵ which decreased its activity at other acidic PHs as well as neutral and basic condition. So, it suggested, for applied this enzyme under others pH conditions, use mutagenesis and/or other methods to increase its pH stability.

کلیدواژه ها

Electrophoresis, Oxidoreductase, Purification

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