Comparative Analysis of Peripheral Alkaline Phytase Protein Structures Expressed in E. coli
سال انتشار: 1394
نوع سند: مقاله ژورنالی
زبان: انگلیسی
مشاهده: 168
فایل این مقاله در 9 صفحه با فرمت PDF قابل دریافت می باشد
- صدور گواهی نمایه سازی
- من نویسنده این مقاله هستم
استخراج به نرم افزارهای پژوهشی:
شناسه ملی سند علمی:
JR_RBMB-4-1_002
تاریخ نمایه سازی: 10 شهریور 1400
چکیده مقاله:
Background: Degradation of phytic acid to inorganic phosphate in domestic animals’ diets requires thermostable phytase. Although Basillus subtilis phytase shows a potential to be degraded phytate complex in high temperature, the enzyme activities and yields need to be increased to make them possible for industrial application.
Methods: The phytase gene from Bacillus subtilis DR۸۸۸۶ was isolated from Dig Rostam hot mineral spring in Iran and cloned into pET۲۱(+) and pET۳۲(+). Expression was induced with ۱.۵ mM IPTG and the proteins were purified.
Results: The recombinant protein affected by thioredoxin (Trx) from pET۳۲a-PhyC was estimated to constitute about ۳۱% of the total soluble protein in the cells; its concentration was ۳.۵ µg/ml, and its maximal phytase activity was ۱۵.۹ U/ml, whereas the recombinant phytase from pET۲۱a-PhyC was estimated to comprise about ۱۹% of the total soluble protein; its concentration was ۲.۲ µg/ml, and its maximal phytase activity was ۶۹ U/ml. The molecular masses of recombinant phytase with and without Trx were about ۶۰ kDa and ۴۲ kDa, respectively. Zymography confirmed that the recombinant enzymes were active. Although the concentration of the alkaline phytase expressed by pET۳۲a was approximately ۵۹% greater than that expressed by pET۲۱, its phytase activity was approximately ۷۷% less.
Conclusion: This study showed that the peripheral gene (Trx) encoded by the pET۳۲a (+) vector are the principal reason for the decrease in recombinant phytase enzyme activity.
کلیدواژه ها:
نویسندگان
Mohammadreza Nassiri
Institute of Biotechnology, Ferdowsi University of Mashhad, Mashhad, Iran - Department of Animal Sciences, College of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran
Hamid Ariannejad
Department of Animal Sciences, College of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran