Scientific Results

Direct, simple derivatization of disulfide bonds in proteins with organic mercury in alkaline medium without any chemical pre-reducing agents

Year: 2014

Authors: Campanella B., Onor M., Ferrari C., D’Ulivo A., Bramanti E.

Autors Affiliation: National Research Council of Italy, C.N.R., Istituto di Chimica dei Composti Organo Metallici-ICCOM- UOS Pisa, Area di Ricerca, Via G. Moruzzi 1, 56124 Pisa, Italy; National Research Council of Italy, C.N.R., Istituto Nazionale di Ottica, INO-UOS Pisa, Area di Ricerca, Via G. Moruzzi 1, 56124 Pisa, Italy

Abstract: In this work we have studied the derivatization of protein disulfide bonds with p-Hydroxymercurybenzoate (. pHMB) in strong alkaline medium without any preliminary reduction. The reaction has been followed by the determination of the protein-. pHMB complex using size exclusion chromatography coupled to a microwave/UV mercury oxidation system for the on-line oxidation of free and protein-complexed pHMB and atomic fluorescence spectrometry (SEC-CVG-AFS) detection. The reaction has been optimized by an experimental design using lysozyme as a model protein and applied to several thiolic proteins. The proposed method reports, for the first time, that it is possible to label 75-100% cysteines of proteins and, thus, to determine thiolic proteins without the need of any reducing step to obtain reduced -SH groups before mercury labelling. We obtained a detection limit of 100 nmol L-1 based on a signal-to-noise ratio of 3 for unbound and complexed pHMB, corresponding to a detection limit of proteins ranged between 3 and 360 nmol L-1, depending on the number of cysteines in the protein sequence.


Volume: 843      Pages from: 1  to: 6

KeyWords: Alkaline medium; Atomic fluorescence spectrometry; Disulfide bond derivatization; P-Hydroxymercuribenzoic acid; Proteins
DOI: 10.1016/j.aca.2014.07.003

Citations: 3
data from “WEB OF SCIENCE” (of Thomson Reuters) are update at: 2022-01-16
References taken from IsiWeb of Knowledge: (subscribers only)
Connecting to view paper tab on IsiWeb: Click here
Connecting to view citations from IsiWeb: Click here

This site uses cookies. If you decide to continue browsing we consider that you accept their use. For more information about cookies and how to delete them please read our Info Policy on cookies use.
Read more