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Probing protein unfolding through monitoring cysteine alkylation by matrix‐assisted laser desorption/ionisation mass spectrometry
Author(s) -
Galvani Marina,
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 - 10.1002/1097-0231(20001030)14:20<1925::aid-rcm113>3.0.co;2-k
Subject(s) - chemistry , mass spectrometry , guanidine , chromatography , cysteine , alkylation , denaturation (fissile materials) , matrix (chemical analysis) , protein mass spectrometry , desorption , matrix assisted laser desorption/ionization , analytical chemistry (journal) , tandem mass spectrometry , organic chemistry , nuclear chemistry , adsorption , enzyme , catalysis
Matrix‐assisted laser desorption/ionisation mass spectrometry was used to monitor interaction between three proteins and two basic Immobiline chemicals (pK 10.3 and pK >12) commonly used in immobilised pH gradients (IPG). For two of the investigated proteins, the observed alkylation channels of the cysteine residues exhibited unmistakable response to their gradual denaturation following treatment with different concentrations (0–8 M) of two commonly used denaturants, urea and guanidine hydrochloride. Our assessment for protein unfolding is based on the number and relative intensity of the alkylation channels, yet the present mass spectrometry data are in good agreement with data based on optical rotatory dispersion, in which another approach was used to assess protein unfolding. Whether the present simple, fast and specific mass spectrometry method can be developed as a probe for monitoring folding/unfolding of cysteine‐containing proteins can only be demonstrated by generating similar data for a larger number of proteins. Copyright © 2000 John Wiley & Sons, Ltd.