INVESTIGATION OF COPPER-ZINC SUPEROXIDE-DISMUTASE SER-137 AND ALA-137 MUTANTS

Ser-and Ala-137 mutants of human copper–zinc superoxide dismutase (Cu,ZnSOD) have been thoroughly characterized in an attempt to understand the subtle effect of the nature of the residue at position 137 on the structure of the copper site and on the activity profile. The results show that the nature of the residue at position 137 determines the presence of water in the active cavity as monitored through water 1H nuclear magnetic relaxation dispersion. Also, the hyperfine shifts experienced by the protons of His-48 in the Cu2Co2 derivative are sensitive to the group at the 137 position. These effects are not detected through electronic and EPR spectroscopies. The activity profiles of Ser-137 and Ala-137 mutants are virtually identical and are very close to that of the Ile-137 mutant at pH < 10. The drop in activity above pH 10 closely parallels that observed in the wild type, at variance with the Ile-137 mutant that shows a marked decrease in activity already below pH 10. The activity profiles definitely show evidence of a pKabetween 6 and 7. The affinity of N3− for the Ser-137 and Ala-137 mutants is very similar to that for the wild type. This is again at variance with the Ile-137 derivative that shows an N3− affinity twice as large. © 1990, American Chemical Society. All rights reserved.