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Probing acrylamide alkylation sites in cysteine‐free proteins by matrix‐assisted laser desorption/ionisation time‐of‐flight
Author(s) -
Bordini Ellenia,
Hamdan Mahmoud,
Righetti Pier Giorgio
Publication year - 2000
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/(sici)1097-0231(20000530)14:10<840::aid-rcm953>3.0.co;2-m
Subject(s) - chemistry , acrylamide , cysteine , monomer , adduct , mass spectrometry , desorption , lysine , chromatography , polymer chemistry , organic chemistry , biochemistry , amino acid , polymer , adsorption , enzyme
It is recognised that gel‐separated proteins can experience a frequent modification provoked by the interaction of unpolymerized acrylamide monomers with the thiol group of cysteine to form a β‐cysteinyl‐ S ‐propionamide adduct. Other groups which have been implicated in this reaction include the hydroxyl group of tyrosine, the ϵ‐amino group of lysine, and the free N‐terminus. In a series of recent publications it has been demonstrated that at pH ∼9.5 and in the presence of cysteine, none of these groups experienced measurable interaction with acrylamide monomers. To emphasise this conclusion we have used matrix‐assisted laser desorption/ionisation with a reflectron time‐of‐flight mass spectrometer to examine a number of cysteine‐free proteins incubated for various intervals with 30 mM acrylamide monomers at pH 9.5. These high resolution data suggest that, for short incubation times (≥1 hour) and in the absence of cysteine, the ϵ‐NH 2 group of lysine is the likely adduction site of acrylamide. Longer incubation times (≥24 hours) with acrylamide monomers rendered the role of Cys as the favourite alkylation site less evident. Copyright © 2000 John Wiley & Sons, Ltd.