Peroxynitrite detoxification by ferryl Mycobacterium leprae truncated hemoglobin O.

During infection, Mycobacterium leprae is faced with the host macrophagic environment limiting the growth of the bacilli. However, (pseudo-)enzymatic detoxification systems, including truncated hemoglobin O (Ml-trHbO), could allow this mycobacterium to persist in vivo. Here, kinetics of peroxynitrite (ONOOH/ONOO(-)) detoxification by ferryl Ml-trHbO (Ml-trHbO-Fe(IV)=O), obtained by treatment with H(2)O(2), is reported. Values of the second-order rate constant for peroxynitrite detoxification by Ml-trHbO-Fe(IV)=O (i.e., of Ml-trHbO-Fe(III) formation; k(on)), at pH 7.2 and 22.0 degrees C, are 1.5x10(4) M(-1) s(-1), and 2.2x10(4) M(-1) s(-1), in the absence of and presence of physiological levels of CO(2) (approximately 1.2x10(-3) M), respectively. Values of k(on) increase on decreasing pH with a pK(a) value of 6.7, this suggests that ONOOH reacts preferentially with Ml-trHbO-Fe(IV)=O. In turn, peroxynitrite acts as an antioxidant of Ml-trHbO-Fe(IV)=O, which could be responsible for the oxidative damage of the mycobacterium. As a whole, Ml-trHbO can undertake within the same cycle H(2)O(2) and peroxynitrite detoxification.

The truncated hemoglobin from Mycobacterium leprae.

Truncated hemoglobins (trHb's) form a family of low molecular weight O2 binding hemoproteins distributed in eubacteria, protozoa, and plants. TrHb's branch in a distinct clade within the hemoglobin (Hb) superfamily. A unique globin gene has recently been identified from the complete genome sequence of Mycobacterium leprae that is predicted to encode a trHb (M. leprae trHbO). Sequence comparison and modelling considerations indicate that monomeric M. leprae trHbO has structural features typical of trHb's, such as 20-40 fewer residues than conventional globin chains, Gly-based sequence consensus motifs, likely assembling into a 2-on-2 alpha-helical sandwich fold, and hydrophobic residues recognized to build up the protein matrix ligand diffusion tunnel. The ferrous heme iron atom of deoxygenated M. leprae trHbO appears to be hexacoordinated, like in Arabidopsis thaliana trHbO-3 (A. thaliana trHbO-3). Accordingly, the value of the second-order rate constant for M. leprae trHbO carbonylation (7.3 x 10(3) M(-1) s(-1)) is similar to that observed for A. thaliana trHbO-3 (1.4 x 10(4) M(-1) s(-1)) and turns out to be lower than that reported for carbon monoxide binding to pentacoordinated Mycobacterium tuberculosis trHbN (6.7 x 10(6) M(-1) s(-1)). The lower reactivity of M. leprae trHbO as compared to M. tuberculosis trHbN might be related to the higher susceptibility of the leprosy bacillus to toxic nitrogen and oxygen species produced by phagocytic cells.