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Título: | Combined use of ESI-MS and UV diode-array detection for localization of disulfide bonds in proteins: application to an α-l-fucosidase of pea |
Autor: | Codina, Ana; Fernández, Irene; Giralt, Ernest; Martínez, Inmaculada; Ludevid, M. Dolors CSIC | Fecha de publicación: | 2001 | Editor: | Ejnar Munksgaard John Wiley & Sons |
Citación: | Journal of Peptide Research 57(6): 473-482 (2001) | Resumen: | A simplified strategy is described for the assignment of disulfide bonds in proteins of medium to high molecular mass (10–30 kDa). The method combines the use of high-performance liquid chromatography coupled to electrospray ionization mass spectrometry (HPLC-ESI-MS) and HPLC with UV diode-array detection (HPLC diode array). The denatured protein is subjected to proteolysis and the peptide mixture is divided into three fractions: (i) underivatized peptides, (ii) ethylpyridylated peptides, and (iii) reduced and ethylpyridylated peptides. The three peptide ensembles are then subjected to chromatographic and spectroscopic analysis. A systematic methodology is described to analyze the large amount of data obtained. The method was applied to the localization of disulfide bonds in α-l-fucosidase from pea. The two disulfide bonds were located between residues Cys64 and Cys109 and between Cys162 and Cys169, while Cys127 was free. | Versión del editor: | https://doi.org/10.1034/j.1399-3011.2001.00859.x | URI: | http://hdl.handle.net/10261/274567 | DOI: | 10.1034/j.1399-3011.2001.00859.x | ISSN: | 1397-002X |
Aparece en las colecciones: | (CID) Artículos |
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