A novel intersubunit communication mechanism in a truncated hemoglobin from Mycobacterium tuberculosis

HbN is a homodimeric hemoglobin from Mycobacterium tuberculosis that belongs to the newly discovered truncated hemoglobin family. The intersubunit communication mechanism in HbN was investigated by ligand replacement experiments, in which the heme-bound CO in the fully CO-saturated dimer was gradually replaced with oxygen, and vice versa. A mixed-ligand species, in which one subunit binds a CO molecule and the other subunit binds an oxygen molecule, was formed under relatively low CO concentrations. The CO-bound subunit in the mixed-ligand species was found in the open conformation, in contrast to an equilibrium mixture of open and closed conformations in the fully CO-saturated derivative. On the other hand, the O₂-bound subunit stays in the closed conformation regardless of the saturation level of CO. These observations suggest that the formation of the H-bonds between the B10 tyrosine and the heme-bound dioxygen in one subunit of the dimer forces its neighboring CO-bound subunit to adopt an open conformation. The new data reported here not only reveal a novel intersubunit communication mechanism but also provide the first experimental evidence for an intermediate ligation state that is critical for a ligand-linked allosteric structural transition.