Recombinant superoxide dismutase from a hyperthermophilic archaeon, Pyrobaculum aerophilium.
Superoxide dismutase (SOD) from the hyperthermophilic archaeon Pyrobaculum aerophilum (a facultative aerobe) has been cloned and expressed in a mesophilic host (Escherichia coli) as a soluble tetrameric apoprotein. The purified apoprotein can be reconstituted with either Mn or Fe by heating the protein with the appropriate metal salt at an elevated temperature (95 degrees C). Both Mn- and Fe-reconstituted P. aerophilum SOD exhibit superoxide dismutase activity, with the Mn-containing enzyme having the higher activity. P. aerophilum SOD is extremely thermostable and the reconstitution with Mn(II) can be performed in an autoclave (122 degrees C, 18 psi). The Mn(III) optical absorption spectrum of Mn-reconstituted P. aerophilum SOD is distinct from that of most other MnSODs and is unchanged upon addition of NaN3. The optical absorption spectrum of Fe-reconstituted P. aerophilum SOD is typical of Fe-substituted MnSODs and authentic FeSOD and exhibits a pH-dependent transition with an effective pKa value higher than that found for Fe-substituted MnSOD from either E. coli or Thermus spp. Amino acid sequence analysis shows that the P. aerophilum SOD is closely related to SODs from other hyperthermophilic archaea (Aeropyrum pernix and Sulfolobus spp.), forming a family of enzymes distinct from the hyperthermophilic bacterial SOD from Aquifex pyrophilus and from mesophilic SODs.
Whittaker
M M
MM
Department of Biochemistry and Molecular Biology, Oregon Graduate Institute of Science and Technology, Portland 97291-1000, USA.
Whittaker
J W
JW
D000595:Amino Acid Sequence; D001665:Binding Sites; D002942:Circular Dichroism; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D004578:Electron Spin Resonance Spectroscopy; D004789:Enzyme Activation; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005822:Genetic Vectors; D006863:Hydrogen-Ion Concentration; D007501:Iron / Q000378:metabolism; D008345:Manganese / Q000378:metabolism; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics*; D017386:Sequence Homology, Amino Acid; D013482:Superoxide Dismutase / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D013696:Temperature; D019624:Thermoproteaceae / Q000201:enzymology*
D017931:DNA Primers; D011994:Recombinant Proteins; D008345:Manganese; D007501:Iron; D013482:Superoxide Dismutase