PURIFICATION END CHARACTERIZATION OF THE CU,ZN SOD FROM ESCHERICHIA-COLI

The periplasmic Cu,Zn superoxide dismutase has been purified to homoge neity by a procedure, which depended upon osmotic shock followed by tw o chromatographic columns. Its subunit weight, determined by electrosp ray ionization mass spectrometry, was found to be 15,737 +/- 1.6. The second derivative ultraviolet spectrum indicated a lack of tryptophan. The amino acid composition as well as a partial N-terminal amino acid sequence is reported. The specific activity was 3700 U/mg and the cor responding copper content was 0.77 atoms Cu/subunit, The enzyme was qu ite unstable and overnight dialysis against EDTA or even prolonged dia lysis against neutral phosphate buffer caused partial loss of activity and of copper and visible precipitation. It is likely that some losse s occurred during the isolation procedure, and if these could have bee n prevented the copper content would have been 1.0 Cu/subunit and the specific activity: would have been 4800 U/mg. It nom appears Likely th at gram negative bacteria will commonly be found to contain a periplas mic Cu,ZnSOD.