کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2817408 1159986 2013 10 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Molecular cloning and catalytic characterization of a recombinant tropine biosynthetic tropinone reductase from Withania coagulans leaf
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی ژنتیک
پیش نمایش صفحه اول مقاله
Molecular cloning and catalytic characterization of a recombinant tropine biosynthetic tropinone reductase from Withania coagulans leaf
چکیده انگلیسی

Tropinone reductases (TRs) are small proteins belonging to the SDR (short chain dehydrogenase/reductase) family of enzymes. TR-I and TR-II catalyze the conversion of tropinone into tropane alcohols (tropine and pseudotropine, respectively). The steps are intermediary enroute to biosynthesis of tropane esters of medicinal importance, hyoscyamine/scopolamine, and calystegins, respectively. Biosynthesis of tropane alkaloids has been proposed to occur in roots. However, in the present report, a tropine forming tropinone reductase (TR-I) cDNA was isolated from the aerial tissue (leaf) of a medicinal plant, Withania coagulans. The ORF was deduced to encode a polypeptide of 29.34 kDa. The complete cDNA (WcTRI) was expressed in E. coli and the recombinant His-tagged protein was purified for functional characterization. The enzyme had a narrow pH range of substantial activity with maxima at 6.6. Relatively superior thermostability of the enzyme (30% retention of activity at 60 °C) was catalytic novelty in consonance with the desert area restricted habitat of the plant. The in vitro reaction kinetics predominantly favoured the forward reaction. The enzyme had wide substrate specificity but did not cover the substrates of other well-known plant SDR related to menthol metabolism. To our knowledge, this pertains to be the first report on any gene and enzyme of secondary metabolism from the commercially and medicinally important vegetable rennet species.


► Withania coagulans leaf cDNA encoding tropine forming tropinone reductase isolated
► The gene (Ws-TR-I) expressed in E. coli and recombinant enzyme homogeneity purified
► Recombinant enzyme characterized for catalytic kinetics and features
► Leaf expression of the gene suggest independent aerial tissue alkaloid biosynthesis
► Genomics and tropane alkaloid biosynthesis description first ever in the plant

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Gene - Volume 516, Issue 2, 10 March 2013, Pages 238–247
نویسندگان
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