A method for terminus proteomics: Selective isolation and labeling of N-terminal peptide from protein through transamination reaction |
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Authors: | Kazuhiro Sonomura Hiroki Kuyama Ei-ichi Matsuo Susumu Tsunasawa Osamu Nishimura |
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Institution: | 1. Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565-0871, Japan;2. Life Science Research Center, Technology Research Laboratory, Shimadzu Corporation, Kyoto 619-0237, Japan |
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Abstract: | A novel method for selectively labeling and isolating N-terminal peptide from protein has been developed. An Nα-amino group of protein was converted to a carbonyl group through transamination reaction and the resulting carbonyl group was modified with O-(4-nitrobenzyl)hydroxylamine (NBHA). After proteolytic digestion using Grifola frondosa metalloendopeptidase (LysN), the modified N-terminal peptide remained unbound in the following treatment using amino-reactive p-phenylenediisothiocyanate (DITC) glass, whereas peptides other than the N-terminal peptide were effectively scavenged from the supernatant solution. The modified N-terminal peptide was thus successfully isolated and sequenced by matrix-assisted laser desorption/ionization tandem mass spectrometry (MALDI-MS/MS) analysis. |
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