The intracellular proteolytic processing of extracellular superoxide dismutase (EC-SOD) is a two-step event

J Biol Chem. 2004 May 21;279(21):22152-7. doi: 10.1074/jbc.M401180200. Epub 2004 Mar 24.


Extracellular superoxide dismutase (EC-SOD) is a tetramer composed of either intact (Trp(1)-Ala(222)) or proteolytically cleaved (Trp(1)-Glu(209)) subunits. The latter form is processed intracellularly before secretion and lacks the C-terminal extracellular matrix (ECM)-binding region ((210)RKKRRRESECKAA(222)-COOH). We have previously suggested that the C-terminal processing of EC-SOD is either a one-step mechanism accomplished by a single intracellular endoproteolytic event cleaving the Glu(209)-Arg(210) peptide bond or a two-step mechanism involving two proteinases (Enghild, J. J., Thogersen, I. B., Oury, T. D., Valnickova, Z., Hojrup, P., and Crapo, J. D. (1999) J. Biol. Chem. 274, 14818-14822). In the latter case, an initial endoproteinase cleavage occurs somewhere in the region between Glu(209) and Glu(216). A carboxypeptidase specific for basic amino acid residues subsequently trims the remaining basic amino acid residues to Glu(209). A naturally occurring mutation of EC-SOD substituting Arg(213) for Gly enabled us to test these hypotheses. The mutation does not prevent proteolysis of the ECM-binding region but prevents a carboxypeptidase B-like enzyme from trimming residues beyond Gly(213). The R213G mutation is located in the ECM-binding region, and individuals carrying this mutation have an increased concentration of EC-SOD in the circulatory system. In this study, we purified the R213G EC-SOD variant from heterozygous or homozygous individuals and determined the C-terminal residue of the processed subunit to be Gly(213). This finding supports the two-step processing mechanism and indicates that the R213G mutation does not disturb the initial endoproteinase cleavage event but perturbs the subsequent trimming of the C terminus.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Aorta / enzymology
  • Carboxypeptidase B / chemistry
  • Carboxypeptidases / chemistry
  • Electrophoresis, Gel, Two-Dimensional
  • Electrophoresis, Polyacrylamide Gel
  • Extracellular Matrix / metabolism
  • Glutamic Acid / chemistry
  • Glycosylation
  • Heterozygote
  • Humans
  • Mass Spectrometry
  • Models, Chemical
  • Molecular Sequence Data
  • Mutation
  • Protein Binding
  • Protein Structure, Tertiary
  • Sequence Homology, Amino Acid
  • Spectrometry, Mass, Electrospray Ionization
  • Superoxide Dismutase / blood
  • Superoxide Dismutase / metabolism*
  • Time Factors


  • Glutamic Acid
  • SOD3 protein, human
  • Superoxide Dismutase
  • Carboxypeptidases
  • Carboxypeptidase B