کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
1214838 1494043 2016 8 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
High-performance thin-layer chromatography as a fast screening tool for phosphorylated peptides
ترجمه فارسی عنوان
کروماتوگرافی لایه نازک با عملکرد بالا به عنوان یک ابزار غربالگری سریع برای پپتید های فسفریلیک شده
موضوعات مرتبط
مهندسی و علوم پایه شیمی شیمی آنالیزی یا شیمی تجزیه
چکیده انگلیسی


• An immunological detection of phosphopeptides after HPTLC was developed.
• Phosphopeptides were detected applying monoclonal antibodies.
• Via HPTLC-MALDI-TOF a simultaneous characterization of the peptides was realized.

This study aimed at developing a rapid chromatographic assay to monitor phosphorylation sites in peptides. For the analysis of nociceptive signal transduction pathways, the detection of phosphorylated proteins/peptides plays a fundamental role. To get further insights in the phosphorylation mechanism of protein kinase C-ε (PKC-ε) and protein kinase A (PKA), potential targets were divided into subsections resulting in peptides that contain only one possible phospho-binding site. The use of high-performance thin-layer chromatography (HPTLC) offers the possibility of a high throughput of samples and the advantage of a quick sample clean-up. A combined strategy of an effect-directed overlay procedure on the TLC plate using specific antibodies (immunostaining, HPTLC-IS) as well as a parallel, direct mass spectrometric methodology by HPTLC-MALDI-TOF-MS was developed. With regard to HPTLC-IS, validation of the data exhibited a lower limit of detection than the traditionally used protein derivatization reagent fluorescamine. Besides the identification of the phosphorylated peptides, a semi-quantitative estimation can be performed with HPTLC-IS.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Journal of Chromatography B - Volume 1008, 1 January 2016, Pages 198–205
نویسندگان
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