Influence of haem environment on the catalytic properties of the tetrathionate reductase TsdA from Campylobacter jejuni

Bifunctional diheme cytochrome c thiosul-fate dehydrogenases/tetrathionate reductases (TsdA) exhibit different catalytic properties depend-ing on the source organism. In the human food-borne intestinal pathogen Campylobacter jejuni , TsdA functions as a tetrathionate reductase en-abling respiration with tetrathionate as an al-ternative electron acceptor. Here, evidence is provided that Cys(138) and Met(255) serve as the sixth ligands of Heme 1 and Heme 2, respec-tively, in the oxidized Cj TsdA wt protein. Re-placement of Cys(138) resulted in a virtually inac-tive enzyme, confirming Heme 1 as the active site heme. Significantly, TsdA variants carrying amino acid exchanges in the vicinity of the elec-tron-transferring Heme 2 (Met(255), Asn(254) and Lys(252)) exhibited markedly altered catalytic properties of the enzyme, showing these residues play a key role in the physiological function of TsdA. The growth phenotypes and tetrathionate reductase activities of a series of Δ tsdA/*tsdA complemen-tation strains constructed in the original host C. jejuni 81116, showed that in vivo , the TsdA variants exhibited the same catalytic properties as the pure, recombinantly produced enzymes. However, variants that catalyzed tetrathionate reduction more effectively than the wild-type enzyme did not allow better growth.