Dr. Grover: We appreciate your concern about the high Km values for H2O2. At a low concentration of H2O2 (submillimolar range), we can also clearly detect hydroxyl radicals. The difference in the adduct concentration generated by the mutants and wild-type is larger at the lower concentration. Although the concentration of peroxide cannot reach the Km value in vivo, the damaged species are produced and accumulate with time. Also, the free radical reaction is a chain reaction: one of its characteristics is that the small difference in initial hydroxyl radical concentration amplifies itself through chain carriers (secondary radicals) to generate a larger difference in the amount of damaged products. The k(cat) values of both mutants are identical to that of wild-type enzyme (ref. 9, 10). Thus the differential in free radical generation, higher with mutant enzymes, is derived from the difference in Km values. This is consistent with free radicals playing a critical role in the progression of FALS, which is an age-dependent disorder. Its onset occurs at the age of about 45-50 years when the protective system against oxidative stress becomes sufficiently weak. Allow me to speculate that low Km values for H2O2 in free radical-generating activity would be detrimental to living systems.