Tetrameric manganese superoxide dismutases from anaerobic Actinomyces.
Superoxide dismutase was isolated from each of the anaerobically grown organisms Actinomyces naeslundii, Actinomyces strain E1S.25D, and Actinomyces odontolyticus. The enzymes were 100,000-110,000 mol wt acidic proteins (pI 4.3-4.6) and contained Mn and Zn, but no detectable Fe. The Mn and Zn content varied with the enzyme source. A. naeslundii superoxide dismutase, specific activity 2200 U/mg, contained 2.3 g atoms Mn and 1.4 g atoms Zn per mole tetramer whereas A. odontolyticus SOD, specific activity 700 U/mg, contained 1.4 g atoms Mn and 1.8 g atoms Zn per mole tetramer. Actinomyces strain E1S.25D, specific activity 1300 U/mg, contained 1.8 g atoms Mn and 1.2 g atoms Zn per mole tetramer. The amino acid compositions of the enzymes were comparable except for arginine, lysine, and tryptophan content. The enzymatic activity of each enzyme was stable in 5 mM H2O2 at 23 degrees C for 2 h. The enzymes were only modestly inhibited by 20 mM NaN3. The enzymatic activity was increased at low ionic strength but was markedly decreased at increased ionic strength with each salt tested except sodium perchlorate, which caused marked inhibition even at low ionic strength. Polyclonal antibodies to A. naeslundii and Actinomyces strain E1S.25D precipitated and inactivated their respective antigens whereas the precipitated A. odontolyticus superoxide dismutase-antibody complex retained virtually full catalytic activity. Immunological studies revealed that the native A. naeslundii and Actinomyces strain E1S.25D MnSODs share common epitopes and cross-reacted with precipitin lines of complete identity in Ouchterlony double diffusion gels. Antibody to the A. odontolyticus enzyme displayed only partial cross-reactivity with superoxide dismutase from the two other Actinomyces. Western blotting of the denatured antigens revealed reactivities of the antibodies that differed only slightly from the results of the Ouchterlony gels.
Barkley
K B
KB
Department of Biochemistry, Virginia Polytechnic Institute, Blacksburg.
Gregory
E M
EM
D000190:Actinomyces / Q000201:enzymology*; D000596:Amino Acids / Q000032:analysis; D000693:Anaerobiosis; D000906:Antibodies; D015153:Blotting, Western; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D005779:Immunodiffusion; D007700:Kinetics; D046911:Macromolecular Substances; D008345:Manganese / Q000032:analysis; D008970:Molecular Weight; D009994:Osmolar Concentration; D013053:Spectrophotometry; D013482:Superoxide Dismutase / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D015032:Zinc / Q000032:analysis
D000596:Amino Acids; D000906:Antibodies; D046911:Macromolecular Substances; D008345:Manganese; D013482:Superoxide Dismutase; D015032:Zinc