کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
10825972 1064692 2013 8 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Workflow for quantitative proteomic analysis of Clostridium acetobutylicum ATCC 824 using iTRAQ tags
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی زیست شیمی
پیش نمایش صفحه اول مقاله
Workflow for quantitative proteomic analysis of Clostridium acetobutylicum ATCC 824 using iTRAQ tags
چکیده انگلیسی
Clostridium acetobutylicum (Cac) is an anaerobic, endospore-forming, Gram-positive bacterium with tremendous promise for use as a biocatalyst for the production of fuels and solvents. Cac proteomic sample preparation for shotgun analysis typically involves a multitude of reagents for harsh lysis conditions and to maintain protein solubility. We describe a protein extraction and preparation method for Cac that is compatible with proteomic shotgun analysis using isobaric labeling approaches. The method is applied to the analysis of Cac grown under butanol stress and labeled using iTRAQ 4-plex reagents. This method relies on the use of calcium carbonate to facilitate lysis by sonication and a commercially available kit to remove detergents prior to labeling. This workflow resulted in the identification and quantitation of 566 unique proteins using ProteinPilot software with a false discovery rate of 0.01% for peptide matches and 0.70% for protein matches. Ninety-five proteins were found to have statistically higher expression levels in butanol-stressed Cac as compared to non-stressed Cac. Sixty-one proteins were found to have statistically lower expression levels in stressed versus non-stressed cells. This method may be applicable to other Gram-positive organisms.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Methods - Volume 61, Issue 3, 15 June 2013, Pages 269-276
نویسندگان
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