کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
1926784 | 1536476 | 2008 | 4 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
p-Dimethylaminocinnamaldehyde derivatization for colorimetric detection and HPLC-UV/vis-MS/MS identification of indoles
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کلمات کلیدی
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
زیست شیمی
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چکیده انگلیسی
Cytochrome P450 2A13 (CYP2A13) is a lung specific enzyme known to activate the potent tobacco procarcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) into two carcinogenic metabolites. CYP2A13 has been crystallized and X-ray diffraction experiments illuminated the structure of this enzyme, but with an unknown ligand present in the enzyme active site. This unknown ligand was suspected to be indole but a selective method had to be developed to differentiate among indole and its metabolites in the protein sample. We successfully modified a microbiological colorimetric assay to spectrophotometrically differentiate between indole and a number of possible indole metabolites in nanomolar concentrations by derivatization with p-dimethylaminocinnamaldehyde (DMACA). Further differentiation of indoles was made by mass spectrometry (HPLC-UV/vis-MS/MS) utilizing the chromophore generated in the DMACA conjugation as a UV signature for HPLC detection. The ligand in the crystallized protein was identified as unsubstituted indole, which facilitated refinement of two alternate conformations in the CYP2A13 crystal structure active site.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Archives of Biochemistry and Biophysics - Volume 475, Issue 1, 1 July 2008, Pages 14-17
Journal: Archives of Biochemistry and Biophysics - Volume 475, Issue 1, 1 July 2008, Pages 14-17
نویسندگان
Patrick R. Porubsky, Emily E. Scott, Todd D. Williams,