A cooperative oxygen-binding hemoglobin from Mycobacterium tuberculosis

Two putative hemoglobin genes, glbN and glbO, were recently discovered in t he complete genome se quence of Mycobacterium tuberculosis H37Rv, Here, we show that the glbN gene encodes a dimeric hemoglobin (HbN) that binds oxyge n cooperatively with very high affinity (P-50 = 0.013 mmHg at 20 degrees C) because of a fast combination (25 mu M-1.s(-1)) and a slow dissociation (0 .2 s(-1)) rate. Resonance Raman spectroscopy and ligand association/dissoci ation kinetic measurements, along with mutagenesis studies, reveal that the stabilization of the bound oxygen is achieved through a tyrosine at the B1 0 position in the distal pocket of the heme with a conformation that is uni que among the globins. Physiological studies performed with Mycobacterium b ovis bacillus Calmette-Guerin demonstrate that the expression of HbN is gre atly enhanced during the stationary phase in aerobic cultures but not under conditions of limited oxygen availability. The results suggest that, physi ologically, the primary role of HbN may be to protect the bacilli against r eactive nitrogen species produced by the host macrophage.