Kinetics of oxidation of aliphatic and aromatic thiols by myeloperoxidase compounds I and II

Myeloperoxidase (MPO) is the most abundant protein in neutrophils and plays a central role in microbial killing and inflammatory tissue damage. Becaus e most of the non-steroidal anti-inflammatory drugs and other drugs contain a thiol group, it is necessary to understand how these substrates are oxid ized by MPO. We have performed transient kinetic measurements to study the oxidation of 14 aliphatic and aromatic mono- and dithiols by the MPO interm ediates, Compound I (k(3)) and Compound II (k(4)), using sequential mixing stopped-flow techniques. The one-electron reduction of Compound I by aromat ic thiols (e.g. methimidazole, 2-mercaptopurine and 6-mercaptopurine) varie d by less than a factor of seven (between 1.39 +/- 0.12 X 10(5) M-1 s(-1) a nd 9.16 +/- 1.63 X 10(5) M-1 s(-1)), whereas reduction by aliphatic thiols, vas demonstrated to depend on their overall net charge and hydrophobic char acter and not on the percentage of thiol deprotonation or redox potential. Cysteamine, cysteine methyl ester, cysteine ethyl ester and alpha-lipoic ac id showed k(3) values comparable to aromatic thiols, whereas a free carboxy group (e.g. cysteine, N-acetylcysteine, glutathione) diminished ks dramati cally. The one-electron reduction of Compound II was far more constrained b y the nature of the substrate. Reduction by methimidazole, 2-mercaptopurine and 6-mercaptopurine showed second-order rate constants (k(4)) of 1.33 +/- 0.08 X 10(5) M-1 s(-1), 5.25 +/- 0.07 X 10(5) M-1 s(-1) and 3.03 +/- 0.07 X 10(3) M-1 s(-1). Even at high concentrations cysteine, penicillamine and glutathione could not reduce Compound II, whereas cysteamine (4.27 +/- 0.05 X 10(3) M-1 s(-1)), cysteine methyl eater (8.14 +/- 0.08 X 10(3) M-1 s(-1) ), cysteine ethyl ester (3.76 +/- 0.17 X 10(3) M-1 s(-1)) and alpha-lipoic acid (4.78 +/- 0.07 X 10(4) M-1 s(-1)) were demonstrated to reduce Compound II and thus could be expected to be oxidized by MPO without co-substrates. (C) 1999 Federation of European Biochemical Societies.