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S-9-8 Physiological function of extracellular-superoxide dismutase and reduction of its heparin-affinity by arginine-213 to glycine mutation
T. Adachi Laboratory of Clinical Pharmaceutics, Gifu Pharmaceutical University, Gifu, 502-8585, Japan

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@Superoxide and other secondary generated reactive oxygen metabolites are continuously generated as normal by-products of cellular metabolism. These oxygen species are highly reactive and can attack any biochemical components in the cells. Normal cells have a number of enzymatic endogenous antioxidant mechanisms, for example, superoxide dismutase (SOD), catalase and glutathions peroxidase. These enzymes quickly eliminate toxic oxygen metabolites under normal conditions. The generation of highly toxic metabolites of oxygen is greatly increased in pathological conditions. When the flux of reactive oxygen metabolites exceeds the capability of the endogenous antioxidant mechanisms, tissue injury ensures. There are three SOD isozymes, copper- and zinc-containing SOD (Cu, Zn-SOD), manganese-containing SOD (Mn-SOD) and extracellular-SOD (EC-SOD). EC-SOD is a secretory, tetrameric glycoprotein with a subunit molecular mass of about 32 kDa. EC-SOD is the major SOD isozyme in plasma, lymph and synovial fluid, whereas the content of EC-SOD in tissues is lower than those of Cu,Zn-SOD and Mn-SOD, which are intracellular enzymes.

@ A prominent feature of EC-SOD is its affinity for heparin, which differentiated it from the other SOD isozymes. In rite vascular system, EC-SOD forms an equilibrium between plasma phase and the endothelial cell surfaces, but occurs primarily in the glycocalyx of cell surfaces and in the connective tissue matrix; this form of the enzyme accounts for over 90% of the EC-SOD. In several pathological situations the rate of superoxide formation by activated neutrophils and xanthine oxidase-based system is increased in the microenvironment created by the adhered neutrophil-endothelial cell surface. The endothelial surface-associated EC-SOD would be an efficient protector against such events in vivo.

@ The carboxyl-terminal position of EC-SOD, especially the cluster of four Arg and two Lys residues at position 210-215, forms an essential part of the heparin-binding domain. Molecular genetic studies of EC-SOD have shown that individuals with 10-fold higher plasma EC-SOD content have a single base substitution generating the exchange of GIy for Arg-213 (R213G). The binding of mutant EC-SOD to endothelial cells was about 50-fold less than that of normal EC-SOD. We found that about 6% of the Japanese population tested have the mutation. The frequency of the identified mutant allele in hemodialysis patients was about twice of that in healthy persons. Although the data may be insufficient, the decrease of the protective capability on the endothelial surface by EC-SOD might accelerate the renal dysfunction.

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