کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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5218516 | 1383330 | 2012 | 6 صفحه PDF | دانلود رایگان |

An assay for the spectrophotometric determination of Baeyer-Villiger monooxygenase (BVMO) activity is described. Baeyer-Villiger oxidation of p-nitroacetophenone generates the corresponding acetate and subsequent hydrolysis of this ester by an esterase or NaOH results in the formation of p-nitrophenolate. This chromophore can be easily quantified spectrophotometrically at 410Â nm. The assay can be performed in a microtiter plate format and is applicable to whole Escherichia coli cells containing recombinant BVMO, crude cell extract as well as using purified enzyme as exemplified for the 4-hydroxyacetophenone monooxygenase (HAPMO) from Pseudomonas putida JD1. Furthermore, the assay was used to identify more active HAPMO variants within enzyme mutant libraries generated by error-prone PCR or site-saturation mutagenesis.
Journal: Tetrahedron - Volume 68, Issue 37, 16 September 2012, Pages 7575-7580