Probing cysteine reactivity in proteins by mass spectrometric EC-tagging.
The on-line electrochemical tagging (EC-tagging) of cysteine residues in proteins during mass spectrometry is studied to probe the cysteine environment. Benzoquinone probes electrogenerated at a microspray electrode react with the thiol functions of the proteins within a microchannel and the products are analyzed by mass spectrometry. The fundamentals of the technique are discussed, with a focus on the kinetic aspects. The EC-tagging efficiency of the cysteine residues in proteins is used to probe their environment. Experiments with unmodified proteins and their chemically reduced forms highlight the strong effect of the cysteine site reactivity on the tagging efficiencies. This study highlights relevant parameters for such on-line electrochemical derivatization/MS detection strategies.
Dayon
Loïc
L
Laboratoire d'Electrochimie Physique et Analytique, Ecole Polytechnique Fédérale de Lausanne, EPFL, CH-1015 Lausanne, Switzerland.
Roussel
Christophe
C
Girault
Hubert H
HH
D000818:Animals; D016227:Benzoquinones / Q000737:chemistry; D002417:Cattle; D003545:Cysteine / Q000737:chemistry*; D004563:Electrochemistry; D007328:Insulin / Q000737:chemistry*; D007700:Kinetics; D007782:Lactoglobulins / Q000737:chemistry*; D008956:Models, Chemical; D008968:Molecular Conformation; D010084:Oxidation-Reduction; D011817:Rabbits; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization
D016227:Benzoquinones; D007328:Insulin; D007782:Lactoglobulins; C004532:quinone; D003545:Cysteine