کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2022111 1069278 2006 7 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Purification and characterization of the second Streptomyces phospholipase A2 refolded from an inclusion body
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی زیست شیمی
پیش نمایش صفحه اول مقاله
Purification and characterization of the second Streptomyces phospholipase A2 refolded from an inclusion body
چکیده انگلیسی

A secreted phospholipase A2 (PLA2) from Streptomyces violaceoruber A-2688, previously identified by us, is the first PLA2 identified in prokaryotes. Genome sequence data of Streptomyces coelicolor A3(2) indicates that the bacterium carries two genes encoding hypothetical PLA2s, which exhibit 100 and 78% identity, respectively, to the S. violaceoruber PLA2. In this study, we named the former and latter proteins as the first and second PLA2s, respectively. When the second PLA2 was expressed in Escherichia coli cells, it formed an inclusion body. The present study demonstrates a method to purify it to homogeneity without the disappearance of the enzymatic activity: the inclusion body was washed with sodium deoxycholate and dissolved in the presence of 2 M urea at pH 12, then refolded by the dilution method. The refolding of enzyme was confirmed by the circular dichroism spectrum. The second PLA2 purified to homogeneity had the same specific activity as that of the S. violaceoruber PLA2 and the yield was approximately 6.8 mg/L culture. The second PLA2 exhibits similar enzymatic properties to the S. violaceoruber PLA2, except that the former enzyme does not utilize phophatidic acid as a substrate. The surface electrostatic potential of the S. coelicolor PLA2 model, which is created by the computer-homology modeling, suggests that the positively charged surface of the enzyme does not affect the substrate specificity.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Protein Expression and Purification - Volume 50, Issue 1, November 2006, Pages 82–88
نویسندگان
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