Reactivity of Sulfenic Acid in Human Serum Albumin

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• 作者：Lucí ; a Turell ; Horacio Botti ; Sebasti&aacute ; n Carballal ; Gerardo Ferrer-Sueta ; José ; M. Souza ; Rosario Dur&aacute ; n ; Bruce A. Freeman ; Rafael Radi ; Beatriz Alvarez
• 刊名：Biochemistry
• 出版年：2008
• 出版时间：January 8, 2008
• 年：2008
• 卷：47
• 期：1
• 页码：358 - 367
• 全文大小：178K
• 年卷期：v.47,no.1(January 8, 2008)
• ISSN：1520-4995

文摘

Sulfenic acid is formed upon oxidation of thiols and is a central intermediate in the redoxmodulation of an increasing number of proteins. Methods for quantifying or even detecting sulfenic acidare scarce. Herein, the reagent 7-chloro-4-nitrobenz-2-oxa-1,3-diazole was determined not to be suitableas a chromophoric probe for sulfenic acid in human serum albumin (HSA-SOH) because of lack ofspecificity. Thionitrobenzoate (TNB) reacted with HSA exposed to hydrogen peroxide, but not control orthiol-blocked HSA. The reaction was biphasic. The first phase was ~20-fold faster than the second phaseand first order in HSA-SOH and TNB (105 ± 11 M^-1 s^-1, 25 C, pH 7.4), allowing quantitative data onHSA-SOH formation and reactivity to be obtained. Exposure of reduced HSA (0.5 mM) to hydrogenperoxide (4 mM, 37 C, 4 min) yielded 0.18 ± 0.02 mol of HSA-SOH per mol of HSA. HSA-SHreacted with hydrogen peroxide at 2.7 ± 0.7 M^-1 s^-1 (37 C, pH 7.4), while HSA-SOH reacted at 0.4± 0.2 M^-1 s^-1, yielding sulfinic acid (HSA-SO₂H), as detected by mass spectrometry. The rate constantsof HSA-SOH with targets of analytical interest such as dimedone and sodium arsenite were determined.HSA-SOH did not react appreciably with the plasma reductants ascorbate or urate, nor with free basicamino acids. In contrast, HSA-SOH reacted rapidly with the plasma thiols cysteine, glutathione,homocysteine, and cysteinylglycine at 21.6 ± 0.2, 2.9 ± 0.5, 9.3 ± 0.9, and 55 ± 3 M^-1 s^-1 (25 C, pH7.4), respectively, supporting a role for HSA-SOH in the formation of mixed disulfides.