کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
1174192 961734 2012 6 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Flavin adenine dinucleotide content of quinone reductase 2: Analysis and optimization for structure–function studies
موضوعات مرتبط
مهندسی و علوم پایه شیمی شیمی آنالیزی یا شیمی تجزیه
پیش نمایش صفحه اول مقاله
Flavin adenine dinucleotide content of quinone reductase 2: Analysis and optimization for structure–function studies
چکیده انگلیسی

Quinone reductase 2 (NQO2) is a broadly expressed enzyme implicated in responses to a number of compounds, including protein kinase inhibitors, resveratrol, and antimalarial drugs. NQO2 includes a flavin adenine dinucleotide (FAD) cofactor, but X-ray crystallographic analysis of human NQO2 expressed in Escherichia coli showed that electron density for the isoalloxazine ring of FAD was weak and there was no electron density for the adenine mononucleotide moiety. Reversed-phase high-performance liquid chromatography (HPLC) of the NQO2 preparation indicated that FAD was not present and only 38% of the protomers contained flavin mononucleotide (FMN), explaining the weak electron density for FAD in the crystallographic analysis. A method for purifying NQO2 and reconstituting with FAD such that the final content approaches 100% occupancy with FAD is presented here. The enzyme prepared in this manner has a high specific activity, and there is strong electron density for the FAD cofactor in the crystal structure. Analysis of NQO2 crystal structures present in the Protein Data Bank indicates that many may have sub-stoichiometric cofactor content and/or contain FMN rather than FAD. This method of purification and reconstitution will help to optimize structural and functional studies of NQO2 and possibly other flavoproteins.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Analytical Biochemistry - Volume 420, Issue 1, 1 January 2012, Pages 84–89
نویسندگان
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