Willem H. Koppenol

Otto Warburg's contributions to current concepts of cancer metabolism

Otto Warburg pioneered quantitative investigations of cancer cell metabolism, as well as photosynthesis and respiration. Warburg and co-workers showed in the 1920s that, under aerobic conditions, tumour tissues metabolize approximately tenfold more glucose to lactate in a given time than normal tissues, a phenomenon known as the Warburg effect. However, this increase in aerobic glycolysis in cancer cells is often erroneously thought to occur instead of mitochondrial respiration and has been misinterpreted as evidence for damage to respiration instead of damage to the regulation of glycolysis. In fact, many cancers exhibit the Warburg effect while retaining mitochondrial respiration. We re-examine Warburgs observations in relation to the current concepts of cancer metabolism as being intimately linked to alterations of mitochondrial DNA, oncogenes and tumour suppressors, and thus readily exploitable for cancer therapy.

THE BASIC CHEMISTRY OF NITROGEN MONOXIDE AND PEROXYNITRITE

After a discussion of the physical chemistry of nitrogen monoxide, such as solubility (1.55 mM at 37 degrees C and an ionic strength of 0.15 M) and diffusion constant (4.8 x 10(-5) cm2s(-1)), several reactions that can acts as sinks are discussed, namely the reaction with dioxygen, with thiols and with superoxide. Of these, the latter reaction leads to a powerful oxidant, peroxynitrite. The thermodynamic and kinetic properties of this molecule are also reviewed.

The Haber-Weiss cycle - 70 years later

Abstract The chain reactions HO• + H2O2 → H2O + O2•- + H+ and O2•- + H+ + H2O2 → O2 + HO• + H2O, commonly known as the Haber-Weiss cycle, were first mentioned by Haber and Willst?tter in 1931. George showed in 1947 that the second reaction is insignificant in comparison to the fast dismutation of superoxide, and this finding appears to have been accepted by Weiss in 1949. In 1970, the Haber-Weiss reaction was revived by Beauchamp and Fridovich to explain the toxicity of superoxide. During the 1970s various groups determined that the rate constant for this reaction is of the order of 1 M-1s-1 or less, which confirmed Georges conclusion. The reaction of superoxide with hydrogen peroxide was dropped from the scheme of oxygen toxicity, and superoxide became the source of hydrogen peroxide, which yields hydroxyl radicals via the Fenton reaction, Fe 2+ + H2O2 → Fe3+ + HO- + HO•. In 1994, Kahn and Kasha resurrected the Haber-Weiss reaction again, but this time the oxygen was believed to be in the singlet (1∆g) state. As toxicity arises not from a Fenton-catalysed Haber-Weiss reaction, but from the Fenton reaction, the Haber-Weiss reaction should not be mentioned anymore.

Chemical characterization of the smallest S-nitrosothiol, HSNO; cellular cross-talk of H2S and S-nitrosothiols.

Dihydrogen sulfide recently emerged as a biological signaling molecule with important physiological roles and significant pharmacological potential. Chemically plausible explanations for its mechanisms of action have remained elusive, however. Here, we report that H2S reacts with S-nitrosothiols to form thionitrous acid (HSNO), the smallest S-nitrosothiol. These results demonstrate that, at the cellular level, HSNO can be metabolized to afford NO+, NO, and NO– species, all of which have distinct physiological consequences of their own. We further show that HSNO can freely diffuse through membranes, facilitating transnitrosation of proteins such as hemoglobin. The data presented in this study explain some of the physiological effects ascribed to H2S, but, more broadly, introduce a new signaling molecule, HSNO, and suggest that it may play a key role in cellular redox regulation.

Mechanism of Reaction of Myeloperoxidase with Nitrite

Myeloperoxidase (MPO) is a major neutrophil protein and may be involved in the nitration of tyrosine residues observed in a wide range of inflammatory diseases that involve neutrophils and macrophage activation. In order to clarify if nitrite could be a physiological substrate of myeloperoxidase, we investigated the reactions of the ferric enzyme and its redox intermediates, compound I and compound II, with nitrite under pre-steady state conditions by using sequential mixing stopped-flow analysis in the pH range 4–8. At 15 °C the rate of formation of the low spin MPO-nitrite complex is (2.5 ± 0.2) × 104 m − 1 s− 1at pH 7 and (2.2 ± 0.7) × 106 m − 1 s− 1at pH 5. The dissociation constant of nitrite bound to the native enzyme is 2.3 ± 0.1 mm at pH 7 and 31.3 ± 0.5 μm at pH 5. Nitrite is oxidized by two one-electron steps in the MPO peroxidase cycle. The second-order rate constant of reduction of compound I to compound II at 15 °C is (2.0 ± 0.2) × 106 m − 1s− 1 at pH 7 and (1.1 ± 0.2) × 107 m − 1s− 1 at pH 5. The rate constant of reduction of compound II to the ferric native enzyme at 15 °C is (5.5 ± 0.1) × 102 m − 1s− 1 at pH 7 and (8.9 ± 1.6) × 104 m − 1s− 1 at pH 5. pH dependence studies suggest that both complex formation between the ferric enzyme and nitrite and nitrite oxidation by compounds I and II are controlled by a residue with a pK a of (4.3 ± 0.3). Protonation of this group (which is most likely the distal histidine) is necessary for optimum nitrite binding and oxidation.

Syntheses of peroxynitrite: to go with the flow or on solid grounds?

Publisher Summary This chapter discusses the synthesis method of peroxynitrite; the best synthesis depends on the application. High concentrations of oxoperoxonitrate(1–), free of nitrate and nitrite, can be achieved with the solid–gas method and the synthesis described by Bohle and coworkers. The ozonization of azide is also convenient. The synthesis must be carried out in a fume hood with a chemical scrubber to scavenge excess ozone. If the reaction is not run to completion, there is contamination with azide. With continued bubbling, nearly all of the azide is oxidized, however some of the oxoperoxonitrate(1–) is also destroyed, and the concentration of nitrite becomes comparable to that of oxoperoxonitrate(1–). Small amounts of azide have major effects on many metalloproteins, and may be a significant problematic contaminant. It will inhibit a number of heme proteins, notably cytochrome- c oxidase of the mitochondrial respiratory chain. The ionic strengths of the quenched-flow and autooxidation syntheses, and to a lesser extent, that of the azide–ozone synthesis, are higher than those that result from dissolving potassium superoxide/oxoperoxonitrate or tetraalkylammonium oxoperoxonitrate in a base of the desired concentration just before an experiment. Another method used is based on the reaction of organic nitrites with hydrogen peroxide.

Human peroxiredoxin 5 is a peroxynitrite reductase.

Peroxiredoxins are an ubiquitous family of peroxidases widely distributed among prokaryotes and eukaryotes. Peroxiredoxin 5, which is the last discovered mammalian member, was previously shown to reduce peroxides with the use of reducing equivalents derived from thioredoxin. We report here that human peroxiredoxin 5 is also a peroxynitrite reductase. Analysis of peroxiredoxin 5 mutants, in which each of the cysteine residues was mutated, suggests that the nucleophilic attack on the O–O bond of peroxynitrite is performed by the N‐terminal peroxidatic Cys47. Moreover, with the use of pulse radiolysis, we show that human peroxiredoxin 5 reduces peroxynitrite with an unequalled high rate constant of (7 ± 3) × 107 M−1 s−1.

THE HABER‐WEISS CYCLE

Abstract— The Haber‐Weiss cycle:

The complex interplay of iron metabolism, reactive oxygen species, and reactive nitrogen species: Insights into the potential of various iron therapies to induce oxidative and nitrosative stress

Production of minute concentrations of superoxide (O2(*-)) and nitrogen monoxide (nitric oxide, NO*) plays important roles in several aspects of cellular signaling and metabolic regulation. However, in an inflammatory environment, the concentrations of these radicals can drastically increase and the antioxidant defenses may become overwhelmed. Thus, biological damage may occur owing to redox imbalance-a condition called oxidative and/or nitrosative stress. A complex interplay exists between iron metabolism, O2(*-), hydrogen peroxide (H2O2), and NO*. Iron is involved in both the formation and the scavenging of these species. Iron deficiency (anemia) (ID(A)) is associated with oxidative stress, but its role in the induction of nitrosative stress is largely unclear. Moreover, oral as well as intravenous (iv) iron preparations used for the treatment of ID(A) may also induce oxidative and/or nitrosative stress. Oral administration of ferrous salts may lead to high transferrin saturation levels and, thus, formation of non-transferrin-bound iron, a potentially toxic form of iron with a propensity to induce oxidative stress. One of the factors that determine the likelihood of oxidative and nitrosative stress induced upon administration of an iv iron complex is the amount of labile (or weakly-bound) iron present in the complex. Stable dextran-based iron complexes used for iv therapy, although they contain only negligible amounts of labile iron, can induce oxidative and/or nitrosative stress through so far unknown mechanisms. In this review, after summarizing the main features of iron metabolism and its complex interplay with O2(*-), H2O2, NO*, and other more reactive compounds derived from these species, the potential of various iron therapies to induce oxidative and nitrosative stress is discussed and possible underlying mechanisms are proposed. Understanding the mechanisms, by which various iron formulations may induce oxidative and nitrosative stress, will help us develop better tolerated and more efficient therapies for various dysfunctions of iron metabolism.

Kinetic study of the reaction of ebselen with peroxynitrite

The second‐order rate constant for the reaction of ebselen with peroxynitrite (ONOO−) is (2.0±0.1) × 106 M−1s−1 at pH ≥ 8 and 25°C, 3–4 orders of magnitude higher than the rate constants observed for cysteine, ascorbate, or methionine. The activation energy is relatively low, 12.8 kJ/mol. This is the fastest reaction of peroxynitrite observed so far. It may allow Secontaining compounds to play a novel role in the defense against peroxynitrite, one of the important reactive species generated during inflammatory processes.