Protein proteolysis and the multi-dimensional electrochromatographic separation of histidine-containing peptide fragments on a chip
| Autor: | Fred E. Regnier, Benjamin E. Slentz, Natalia A. Penner |
|---|---|
| Rok vydání: | 2003 |
| Předmět: |
chemistry.chemical_classification
Capillary electrochromatography Chromatography Monolithic HPLC column medicine.diagnostic_test biology Hydrolysis Proteolysis Organic Chemistry Proteins Peptide General Medicine Reversed-phase chromatography Biochemistry Peptide Fragments Analytical Chemistry chemistry Affinity chromatography Electrochromatography medicine biology.protein Histidine Bovine serum albumin |
| Zdroj: | Journal of Chromatography A. 984:97-107 |
| ISSN: | 0021-9673 |
| Popis: | This paper reports a system for three-dimensional electrochromatography in a chip format. The steps involved included trypsin digestion, copper(II)-immobilized metal affinity chromatography [Cu(II)-IMAC] selection of histidine-containing peptides, and reversed-phase capillary electrochromatography of the selected peptides. Trypsin digestion and affinity chromatography were achieved in particle-based columns with a microfabricated frit whereas reversed-phase separations were executed on a column of collocated monolithic support structures. Column frits were designed to maintain constant cross sectional area and path length in all channels and to retain particles down to a size of 3 microm. Cu(II)-IMAC selection of histidine-containing peptides from standard peptide mixtures and protein digests followed by reversed-phase chromatography of the selected peptides was demonstrated in the electrochromatography mode. The possibility to run a comprehensive proteomic analysis by combining trypsin digestion, affinity selection, and a reversed-phase separation on chips was shown using fluorescein isothiocyanate-labeled bovine serum albumin as an example. |
| Databáze: | OpenAIRE |
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