title: Mn-superoxide dismutase from the halophilic halotolerant bacterium Ba1--isolation and active site spectroscopic studies.
abstract: The superoxide-dismutase (SOD) enzyme, isolated from the halophilic halotolerant bacterium Ba1, was found to be a dimer with a molecular weight of 40 kD and a subunit weight of 23.5 kD. The partial N-terminal sequence showed significant homology to SODs isolated from various sources. Metal analysis showed that SOD from Ba1 contains manganese and iron with the following stoichiometries: 0.9 +/- 0.4 Mn/dimer and 0.6 +/- 0.2 Fe/dimer. Two bands were obtained by isoelectric-focusing, at pI of 4.45 and at 4.40. Native SOD from Ba1 at room temperature was ESR silent. An ESR spectrum of hydrated Mn(II) was obtained from denaturated enzyme. Native enzyme cooled to 97 K showed an ESR spectrum identified as being due to Fe(III). The spectrum was pH-independent. SOD from Ba1 was not inactivated by H2O2. On the basis of these observations, SOD from Ba1 was characterized as MnSOD. The excitation fluorescence spectrum of SOD from Ba1 showed four main peaks in the visible region. The effects on the spectra of KSCN, NaN3, NaF, and ascorbate were examined. Measurements of H2(17)O-nmr relaxation times T1 and T2, for solutions containing E. coli MnSOD and FeSOD, showed no paramagnetic contribution. These results support the assumption that the water molecule at the active site is strongly bound.
authors.affiliation: Department of Chemistry, Technion-Israel Institute of Technology, Haifa, Israel.
authors.forename: Y
authors.initials: Y
authors.lastname: Yorkovsky
authors.forename: B L
authors.initials: BL
authors.lastname: Silver
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D004926:Escherichia coli / Q000201:enzymology*; D006088:Gram-Negative Aerobic Bacteria / Q000201:enzymology* / Q000378:metabolism; D016014:Linear Models; D008969:Molecular Sequence Data; D012680:Sensitivity and Specificity; D017386:Sequence Homology, Amino Acid; D012965:Sodium Chloride / Q000378:metabolism*; D013045:Species Specificity; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry; D013482:Superoxide Dismutase / Q000737:chemistry / Q000302:isolation & purification*
chemical_list: D012965:Sodium Chloride; D013482:Superoxide Dismutase