Biophysical and structural characterization of the putative nickel chaperone CooT from Carboxydothermus hydrogenoformans.

Abstract

Carboxydothermus hydrogenoformans is a model microorganism for the study of [NiFe]-CODH, akey enzyme of carbon cycle in anaerobic microorganisms. The enzyme possesses a unique active site (C-cluster), constituted of a distorted [NiFe3S4] cubane linked to a mononuclear Fe(II) center. Both the biogenesis of the C-cluster and the activation of CODH by nickel insertion remain unclear. Among the three accessory proteins thought to play a role in this latter step (CooC, CooJ, and CooT), CooT is identified as a nickel chaperone involved in CODH maturation in Rhodospirillum rubrum. Here, we structurally and biophysically characterized a putative CooT protein present in C. hydrogenoformans (pChCooT). Despite the low sequence homologies between CooT from R. rubrum (RrCooT) and pChCooT (19% sequence identity), the two proteins share several similarities, such as their overall structure and a solvent-exposed Ni(II)-binding site at the dimer interface. Moreover, the X-ray structure of pChCooT reveals the proximity between the histidine 55, a potential nickel-coordinating residue, and the cysteine 2, a highly conserved key residue in Ni(II)-binding.