AN ARGININE-213 TO GLYCINE MUTATION IN HUMAN EXTRACELLULAR-SUPEROXIDEDISMUTASE REDUCES SUSCEPTIBILITY TO TRYPSIN-LIKE PROTEINASES

Molecular genetic studies of extracellular-superoxide dismutase (EC-SO D) have shown that individuals with high serum EC-SOD content have a s ingle base substitution generating the exchange of glycine for arginin e-213 (R213G) in the heparin-binding domain of this enzyme [Sandstrom, J. et al, (1994) J. Biol. Chem. 269, 19163-19166], which causes the i mpairment of its binding ability to endothelial cell surface [Adachi, T. et al. (1996) Biochem. J. 313, 235-239]. Serum EC-SOD in healthy in dividuals without the above mutation is heterogeneous with regard to h eparin affinity and consists of five fractions, forms (I) to (V), of w hich (IV) and (V) are the main fractions with high affinity for hepari n [Adachi, T. et al. (1995) J. Biochem. 117, 586-590], whereas the maj or fraction in hemodialysis patients was serum EC-SOD form (I), which is thought to be the proteolytic truncated form. On the other hand, se rum EC-SOD in both healthy individuals and hemodialysis patients with the R213G mutation consisted mainly of the high heparin-affinity type. This observation suggests that the susceptibility of EC-SOD to protei nases is reduced by the RZ13G mutation, The affinity of normal EC-SOD (n-EC-SOD) for heparin decreased by the treatment with trypsin, accomp anied by a reduction in the molecular mass. The IC50 of trypsin for th e heparin affinity of R213G mutant EC-SOD (m-EC-SOD) was 0.15 mu g/ml, fivefold that for n-EC-SOD. Heparin affinity of n-EC-SOD was again mo re susceptible to neutrophils than that of m-EC-SOD. These results sug gested that m-EC-SOD is more resistant to trypsin and neutrophil-relea se trypsin-like proteinases than n-EC-SOD, which, causes the heparin a ffinity of serum EC-SOD to differ in individuals with and without the R213G mutation.