GENERATION OF H2O2 IN BIOMEMBRANES T. Ramasarma Biochemica et Biophysica Acta, 694 (1982) 69-93 From pages 70-71 I. Introduction Knowledge of the generation of H2O2 in cellular oxidations has existed for many years. It has ben assumed that H2O2 is toxic to cells and the presence of catalase is indicative of a detoxication mechanism. Other radicals of oxygen were recently recognized to be more potent destructive agents of biological material than H2O2. Also catalase and other peroxidases utilize H2O2 in some cellular oxidation processes leading to several important metabolites. Thus, the generation of H2O2 in cellular processes seems to be purposeful and H2O2 can not be dismissed as a mere undesirable byproduct. Biological formation of H2O2 is not limited to the previously known flavoprotiens and some copper enzymes, but other redox systems, particulrly heme and non-heme iron proteins, are now found to undergo auto-oxidation yeilding H2O2. The capacity for generation of H2O2 is now found to be widespread in a variety of organisms and in the organelles of the cells. The reduction of oxygen to H2O by mitochondrial cytochrome oxidase beingthe predominant oxygen-utilizing reaction had overshadowed the importance of the quantitatively minor pathways. Under aerobic conditions generation of H2O2 by a variety of biomembranes has now been found to be a physiological event interlinked with phenomena such as phagocytosis, transport processes and thermogenesis in some as yet unidentified way. The underlying mechanisms of of these processes seem to involve generation and utilization of H2O2 in mitichondria, microsomes, peroxisomes or plasma membranes. This review gives an account of the potential of the biomembranes to generate H2O2 and its implication in the cellular processes. I A. Steps in the reduction of oxygen Molecular oxygen has two unpaired electrons each of which goes into separate antibonding pi-orbitals with parallel spins giving the molecule the stability and paramagnetic property in the ground state. The reductions of O2 by addition of one ,two and four electrons lead to formation of superoxide anion(O2-), H2O2 and H2O, respectively. (a) O2 >> &1e->> O2- >> &1e->> O2-- (b) O2 >> H+ &1e->> HO2- >> H+&1e->> H2O2 >> H+&1e->> H2O & HO H2O & HO >> H+&1e->> 2H2O Only two electrons can be accomodated by each oxygen atom. The antibonding orbital of molecular oxygen recieves the added electrons and each addition weakens and increases the length of the O to O bond, from 1.274 angstrom in O2 to 1.480 angstrom in H2O2, leading to rupture [1]. The two electron reduction of oxygen dierectly to H2O2 is restricted by symmetry considerations [2] that can be overcome by binding of O2 to the electron donor and consequent pertubution of the molecular orbitals. (c) O2 & 2H+ & 2e- >> H2O2 (d) O2 & 1e- >> O2- (e) O2- & O2- & 2H+ >> H2O2 & O2 The flavoprotein oxidases appear to follow this type of direct two electron reduction process (reaction c) with no intermediate step [3]. Other H2O2 generating systems seem to use one electron reductions forming superoxide anions (O2-)(reaction d)[4] two of which then dismutate yielding a molecule each of H2O2 and O2 either spontaneously or catalyzed by the enzyme superoxide dismutase (reaction e)[5]. The flavoprotein dehydrogenases and possibly the theiron protein generating H2O2 seem to adopt this mechanism ans are mostly membrane- localized. It is now found that superoxide formation is a property shared by large number of redox components. In view of the ubiquitous nature of superoxide dismutase and easy nonenzymatic dismutation of superoxide, generation of H2O2 accompanying oxidation of these redox components with molecular oxygen becomes equally widespread. References 1 Samuel, D. and Steckel, F. (1974) in Molecular Oxygen in Biology - Topics in Molecular Oxygen Research (Hayashi,O., ed.), pp. 1-27, North-Holland Pbl. Co, Amsterdam 2 Taube, H. (1965) J. Gen. Physiol. 49, 29-35 3 Massey, V., Strickland, S., Mayhew, S.G., Howell,L.G., Engle, P.C., Mathews, R.g., Schuman, M. and sullivan, P.A. (1969) Biochem. Biophys. Res. Commun. 36,891-897 4 Fridovich, I. and Handler, P. (1961) J. Biol. Chem. 236,1836-1840 5 McCord, J.M. and Fridovich, I. (1969) J. Biol. Chem. 244,6049-6055