Requirements for superoxide-dependent tyrosine hydroperoxide formation in peptides

TY - JOUR

T1 - Requirements for superoxide-dependent tyrosine hydroperoxide formation in peptides

AU - Winterbourn, Christine C

AU - Parsons-Mair, Helena N

AU - Gebicki, Silvia

AU - Gebicki, Janusz M

AU - Davies, Michael Jonathan

PY - 2004/7/1

Y1 - 2004/7/1

N2 - Superoxide reacts rapidly with other radicals, but these reactions have received little attention in the context of oxidative stress. For tyrosyl radicals, reaction with superoxide is 3-fold faster than dimerization, and forms the addition product tyrosine hydroperoxide. We have explored structural requirements for hydroperoxide formation using tyrosine analogues and di- and tri-peptides. Superoxide and phenoxyl radicals were generated using xanthine oxidase, peroxidase and the respective tyrosine derivative, or by gamma-radiation. Peroxides were measured using FeSO4/Xylenol Orange. Tyrosine and tyramine formed stable hydroperoxides, but N-acetyltyrosine and p-hydroxyphenylacetic acid did not, demonstrating a requirement for a free amino group. Using [14C]tyrosine, the hydroperoxide and dityrosine were formed at a molar ratio of 1.8:1. Studies with pre-formed hydroperoxides, and measurements of substrate losses, indicated that, in the absence of a free amino group, reaction with superoxide resulted primarily in restitution of the parent compound. With dipeptides, hydroperoxides were formed only on N-terminal tyrosines. However, adjacent lysines promoted hydroperoxide formation, as did addition of free lysine or ethanolamine. Results are compatible with a mechanism [d'Alessandro, Bianchi, Fang, Jin, Schuchmann and von Sonntag (2000) J. Chem. Soc. Perkin Trans. II, 1862-1867] in which the phenoxyl radicals react initially with superoxide by addition, and the intermediate formed either releases oxygen to regenerate the parent compound or is converted into a hydroperoxide. Amino groups favour hydroperoxide formation through Michael addition to the tyrosyl ring. These studies indicate that tyrosyl hydroperoxides should be formed in proteins where there is a basic molecular environment. The contribution of these radical reactions to oxidative stress warrants further investigation.

AB - Superoxide reacts rapidly with other radicals, but these reactions have received little attention in the context of oxidative stress. For tyrosyl radicals, reaction with superoxide is 3-fold faster than dimerization, and forms the addition product tyrosine hydroperoxide. We have explored structural requirements for hydroperoxide formation using tyrosine analogues and di- and tri-peptides. Superoxide and phenoxyl radicals were generated using xanthine oxidase, peroxidase and the respective tyrosine derivative, or by gamma-radiation. Peroxides were measured using FeSO4/Xylenol Orange. Tyrosine and tyramine formed stable hydroperoxides, but N-acetyltyrosine and p-hydroxyphenylacetic acid did not, demonstrating a requirement for a free amino group. Using [14C]tyrosine, the hydroperoxide and dityrosine were formed at a molar ratio of 1.8:1. Studies with pre-formed hydroperoxides, and measurements of substrate losses, indicated that, in the absence of a free amino group, reaction with superoxide resulted primarily in restitution of the parent compound. With dipeptides, hydroperoxides were formed only on N-terminal tyrosines. However, adjacent lysines promoted hydroperoxide formation, as did addition of free lysine or ethanolamine. Results are compatible with a mechanism [d'Alessandro, Bianchi, Fang, Jin, Schuchmann and von Sonntag (2000) J. Chem. Soc. Perkin Trans. II, 1862-1867] in which the phenoxyl radicals react initially with superoxide by addition, and the intermediate formed either releases oxygen to regenerate the parent compound or is converted into a hydroperoxide. Amino groups favour hydroperoxide formation through Michael addition to the tyrosyl ring. These studies indicate that tyrosyl hydroperoxides should be formed in proteins where there is a basic molecular environment. The contribution of these radical reactions to oxidative stress warrants further investigation.

KW - Amines

KW - Dimerization

KW - Free Radicals

KW - Hydrogen Peroxide

KW - Models, Chemical

KW - Peptides

KW - Superoxides

KW - Tyrosine

U2 - 10.1042/BJ20040259

DO - 10.1042/BJ20040259

M3 - Journal article

C2 - 15025556

SN - 0264-6021

VL - 381

SP - 241

EP - 248

JO - Biochemical Journal

JF - Biochemical Journal

IS - Pt 1

ER -