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'); Reports of Biochemistry and Molecular Biology rbmb.net Basic Sciences http://rbmb.net 1 admin 2322-3480 2322-3480 10.61882/rbmb en jalali 1394 7 1 gregorian 2015 10 1 4 1 online 1 fulltext
en Comparative Analysis of Peripheral Alkaline Phytase Protein Structures Expressed in E. coli بیوشیمی Biochemistry <p style="text-align: justify;"><em><strong>Background: </strong></em>Degradation of phytic acid to inorganic phosphate in domestic animals&rsquo; diets requires thermostable phytase. Although <em>Basillus subtilis</em> 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.</p> <p style="text-align: justify;"><em><strong>Methods:</strong></em> The phytase gene from <em>Bacillus subtilis</em> DR8886 was isolated from Dig Rostam hot mineral spring in Iran and cloned into pET21(+) and pET32(+). Expression was induced with 1.5 mM IPTG and the proteins were purified.</p> <p style="text-align: justify;"><em><strong>Results:</strong></em> The recombinant protein affected by thioredoxin (Trx) from pET32a-PhyC was estimated to constitute about 31% of the total soluble protein in the cells; its concentration was 3.5 &micro;g/ml, and its maximal phytase activity was 15.9 U/ml, whereas the recombinant phytase from pET21a-PhyC was estimated to comprise about 19% of the total soluble protein; its concentration was 2.2 &micro;g/ml, and its maximal phytase activity was 69 U/ml. The molecular masses of recombinant phytase with and without Trx were about 60 kDa and 42 kDa, respectively. Zymography confirmed that the recombinant enzymes were active. Although the concentration of the alkaline phytase expressed by pET32a was approximately 59% greater than that expressed by pET21, its phytase activity was approximately 77% less.</p> <p style="text-align: justify;"><em><strong>Conclusion:</strong></em> This study showed that the peripheral gene (Trx) encoded by the pET32a (+) vector are the principal reason for the decrease in recombinant phytase enzyme activity.</p> Bacillus subtilis DR8806, PhyC Gene, Thioredoxin 10 18 http://rbmb.net/browse.php?a_code=A-10-1-45&slc_lang=en&sid=1 Mohammadreza Nassiri Nassiryr@um.ac.ir 10031947532846006085 10031947532846006085 Yes Institute of Biotechnology, Ferdowsi University of Mashhad, Mashhad, Iran - Department of Animal Sciences, College of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran Hamid Ariannejad 10031947532846006086 10031947532846006086 No Department of Animal Sciences, College of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran