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- W2364507628 abstract "Objective: To express the rabbit NADP(H)-dependent retinol dehydrogenase/reductase(NRDR)in prokaryotic expression system functionally,and to identify its enzyme activity.Methods: The total NRDR sequence was cloned from the rabbit liver and the coding region of NRDR was constructed to the Gateway-based expression vector(pDEST 17),which was transformed into the Escherichia coli(E.coli)(BL21-AI)for the protein expression.After sonication and centrifugation of the bacteria,the soluble NRDR in supernatant was purified by affinity chromatography.Furthermore,the kinetic parameters of NRDR were determined by high performance liquid chromatography(HPLC).Results: The expression vector with the desired gene(768 bp)was constructed,and then,the NRDR protein,which was harvested at the optimal time point(4 hours after induction),was successfully expressed with a 41.4% expression level.Moreover,the homogeneous enzyme was obtained by a one-step affinity chromatography,with purity up to 97.2% and specific activity 64 fold enhanced.The values of the Km and the Vmax were(4.55±0.33)μmol/L and(0.307±0.065)μmol/(min·mg),respectively.Conclusion: The functional rabbit NRDR can be expressed effectively in BL21-AI by pDEST 17." @default.
- W2364507628 created "2016-06-24" @default.
- W2364507628 creator A5077627564 @default.
- W2364507628 date "2007-01-01" @default.
- W2364507628 modified "2023-09-23" @default.
- W2364507628 title "Functional Expression,Purification and Activity Identification of Rabbit NADP(H)-Dependent Retinol Dehydrogenase/Reductase" @default.
- W2364507628 hasPublicationYear "2007" @default.
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