Mycobacteria secrete the enzyme superoxide dismutase (SOD) which forms part of their resistance to oxidative killing by host phagocytic cells. Professor Douglas Young at St Mary's Medical School (London) has cloned and expressed this enzyme which subsequently crystallised and yielded diffraction data to 2.0 Angstroms. The structure has been refined to an R-factor of 0.167 and shows that this enzyme consists of two domains with the catalytic ferric ion located between. The N-terminal domain consists of an extended region followed by two long antiparallel helical segments which form an arm-like structure. The C-terminal domain consists of four more helices and has a three-stranded beta-sheet inserted between the enzyme's 4th and 5th helices.
The above figure shows the tertiary structure of the M. tuberculosis SOD with the catalytic iron (green sphere) and its histidine and solvent ligands (deep blue) together with the coordinating aspartate (red).
As for other enzymes in the Fe- or Mn-dependent SOD family, the catalytic metal ion is coordinated by three invariant histidines, one aspartate and a solvent ligand which is probably a hydroxide ion. This ligand interacts with an enzyme residue which is either His or Gln. There may be a correlation between the nature of this residue and the metal ion specificity of the enzyme as Fe-dependent SODs generally have His at this position whereas Mn-SODs frequently have Gln.
The M. tuberculosis SOD is a compact tetramer, unlike some SODs which are dimeric. The quaternary interactions between the dimers within the tetrameric enzymes vary greatly although the dimers themselves are structurally conserved. The dimer-dimer interactions in tetrameric SODs are dominated by two regions namely the helical arm in the N-terminal domain and the loop connecting the two outer strands of the beta-sheet in the other domain (see above figure). These two regions differ greatly in both sequence and structure between enzymes in this family allowing a range of quaternary structures to be adopted. In the dimeric SODs the helical arm is bent over and interacts with the loop in the other domain and may therefore prevent tetramer formation.
We are currently analysing mutants of this enzyme designed to investigate the determinants of its metal ion specificity and quaternary structure. Crystallisation trials of other M. tuberculosis antigens are underway.
Researchers: Mohammed Badasso, Jane Jackson, Jon Cooper & Steve Wood.
X-ray structure analysis of the iron-dependent superoxide dismutase from Mycobacterium tuberculosis at 2.0 Angstroms resolution reveals novel dimer-dimer interactions.
J.B.Cooper, K.McIntyre, M.O.Badasso, S.P.Wood, Y.Zhang, T.R.Garbe and D.Young. J.Mol.Biol. (1995) in press.