On-line cysteine modification for protein analysis: new probes for electrochemical tagging nanospray mass spectrometry

Generation of mass tags by the inherent electrochemistry of electrospray for protein mass spectrometry

“…To obtain more accurate values of the different kinetic constants k, the evolutions of the I p /I p 0 ratios versus the CVs scan rate were fitted to the experimental data, as shown in Figure 5b for hydroquinone 1c. The value of k was found to increase from 1a to 1d (see Table 1) [66,68]. This result could easily be explained by the fact that the coupling reaction involves a nucleophile (L-cysteine 3a=): in that case, the presence of an electron donor group such as CH 3 or CH 3 O increases the electronic density on the benzoquinone ring and is expected to decrease its reactivity toward L-cysteine addition.…”

“…Hydroquinones 1a-d/benzoquinones 2a-d proved to be stable at the cyclic voltammetry (CV) time scale ( Figure 4, thin lines). The long-term stability of these compounds was checked by UV and 1 H NMR studies [68]. From an electrochemical point of view, the hydroquinone CVs are irreversible, exhibiting a slow electrode kinetic process.…”

“…From an electrochemical point of view, the hydroquinone CVs are irreversible, exhibiting a slow electrode kinetic process. Moreover, CV of pure L-cysteine 3a= revealed the nonelectroactive character of this compound in the experimental conditions [66,68]. The electrochemical tagging of L-cysteine by hydroquinones 1a-d was then investigated by cyclic voltammetry in the MS spray medium: the CVs obtained with pure hydroquinones (oxidation current intensity I p 0 ) were compared with those of the same hydroquinones in the presence of L-cysteine (oxidation current intensity I p ).…”

“…Based on recent works on the electrochemical detection of sulfides [60,61], thiols [62], and cysteine [63,64] with quinone derivatives, we focused on the potential use of electrogenerated benzoquinones (coming from the oxidation of hydroquinones) able to react specifically with cysteine residues during electrospray mass spectrometric measurements. We present below a review of the different steps of the development of on-line electrochemical probes for cysteine mass tagging, from the proof of principle [65] to the elucidation of the tagging reaction mechanism [66], characterization of the microspray design influence on the tagging process [67], search for an optimal electrochemical probe [68], and final application to protein identification [69].…”

Electrochemical multi-tagging of cysteinyl peptides during microspray mass spectrometry: numerical simulation of consecutive reactions in a microchannel

“…The addition rate constants are assumed to be equal to those corresponding to the addition of L-cysteine on 1,4-benzoquinone and methoxycarbonyl-1,4-benzoquinone (210 and 5000 M À1 s À1 , respectively, in methanol-water-acetic acid 50 : 49 : 1). 26,27 For simplification, the diffusion coefficient of all the quinone probes is taken to be equal to 3.5 Â 10 À10 m 2 s À1 , which in fact corresponds to the diffusion coefficient of methoxycarbonyl-1,4-hydroquinone in methanol-water-acetic acid 50 : 49 : 1. 28 The mean diffusion coefficient of the target biomolecule P was chosen to be 1 Â 10 À10 m 2 s À1 .…”