Spectroscopic and redox properties of a CooA homologue from Carboxydothermus hydrogenoformans

Abstract

CooA is a CO-sensing transcriptional activator that contains a b-type heme as the active site for sensing its physiological effector, CO. In this study, the spectroscopic and redox properties of a new CooA homologue from Carboxydothermus hydrogenoformans (Ch-CooA) were studied. Spectroscopic and mutagenesis studies revealed that His-82 and the N-terminal α-amino group were the axial ligands of the Fe(III) and Fe(II) hemes in Ch-CooA and that the N-terminal α-amino group was replaced by CO upon CO binding. Two neutral ligands, His-82 and the N-terminal α-amino group, are coordinated to the Fe(III) heme in Ch-CooA, whereas two negatively charged ligands, a thiolate from Cys-75 and the nitrogen atom of the N-terminal Pro, are the axial ligands of the Fe(III) heme in Rr-CooA. The difference in the coordination structure of the Fe(III) heme resulted in a large positive shift of redox potentials of Ch-CooA compared with Rr-CooA. Comparing the properties of Ch-CooA and Rr-CooA demonstrates that the essential elements for CooA function will be: (i) the heme is six-coordinate in the Fe(III), Fe(II), and Fe(II)-CO forms; (ii) the N-terminal is coordinated to the heme as an axial ligand, and (iii) CO replaces the N-terminal bound to the heme upon CO binding.