Quantification of cysteine residues following oxidation to cysteic acid in the presence of sodium azide

Analytical Biochemistry
M MannebergM Fountoulakis

Abstract

Quantification of cysteines by amino acid composition analysis is inaccurate because of decomposition of these residues during protein hydrolysis. Cysteine (and cystine) residues are oxidized to cysteic acid following hydrochloric acid hydrolysis in the presence of sodium azide. Using selected native and recombinant proteins, containing different numbers of cysteine residues, we investigated the conditions for the quantitative oxidation of cysteines to cysteic acid in the presence of sodium azide. Protein hydrolysis with hydrochloric acid in the presence of 0.20% sodium azide resulted in 87-100% oxidation of the cysteines to cysteic acid which was easily quantified. The results were highly reproducible so that the azide-induced oxidation can be used as a general method to determine cysteine residues in a given protein. The sodium azide-dependent oxidation is superior to oxidation with performic acid because (i) it can be performed in solution not requiring protein lyophilization and in approximately half of the time; (ii) it delivers slightly higher yields of cysteic acid; and (iii) it does not affect tyrosine residues, which can be modified during the performic acid treatment.

Citations

Jul 27, 1999·Protein Expression and Purification·S J Hong, K W Kang
Nov 18, 2020·Rapid Communications in Mass Spectrometry : RCM·Alexandra A PhillipsAlex L Sessions
Jan 23, 1999·Journal of Chromatography. a·M Fountoulakis, H W Lahm
Apr 7, 1998·Journal of Chromatography. a·M WeissM Fountoulakis
Oct 1, 1996·Applied and Environmental Microbiology·U SauerJ E Bailey
Jan 10, 2014·Journal of Agricultural and Food Chemistry·Shane M RutherfurdPaul J Moughan

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