Reaction of Mycobacterium tuberculosis cytochrome P450 enzymes with nitric oxide.
During the initial growth infection stage of Mycobacterium tuberculosis (Mtb), (*)NO produced by host macrophages inhibits heme-containing terminal cytochrome oxidases, inactivates iron/sulfur proteins, and promotes entry into latency. Here we evaluate the potential of (*)NO as an inhibitor of Mtb cytochrome P450 enzymes, as represented by CYP130, CYP51, and the two previously uncharacterized enzymes CYP125 and CYP142. Using UV-visible absorption, resonance Raman, and stopped-flow spectroscopy, we investigated the reactions of (*)NO with these heme proteins in their ferric resting form. (*)NO coordinates tightly to CYP125 and CYP142 (submicromolar) and with a lower affinity (micromolar) to CYP130 and CYP51. Anaerobic reduction of the ferric-NO species with sodium dithionite led to the formation of two spectrally distinct classes of five-coordinate ferrous-NO complexes. Exposure of these species to O(2) revealed that the ferrous-NO forms of CYP125 and CYP142 are labile and convert back to the ferric state within a few minutes, whereas ferrous CYP130 and CYP51 bind (*)NO almost irreversibly. This work clearly indicates that, at physiological concentrations (approximately 1 microM), (*)NO would impair the activity of CYP130 and CYP51, whereas CYP125 and CYP142 are more resistant. Selective P450 inhibition may contribute to the inhibitory effects of (*)NO on Mtb growth.
Ouellet
Hugues
H
Department of Pharmaceutical Chemistry, University of California at San Francisco, San Francisco, California 94158-2517, USA.
Lang
Jérôme
J
Couture
Manon
M
Ortiz de Montellano
Paul R
PR
D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism*; D002417:Cattle; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D005290:Ferric Compounds / Q000378:metabolism; D006736:Horses; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000254:growth & development; D009569:Nitric Oxide / Q000096:biosynthesis / Q000378:metabolism*; D011485:Protein Binding; D026361:Reactive Nitrogen Species / Q000096:biosynthesis / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism
D001426:Bacterial Proteins; C527931:CYP130 protein, M tuberculosis; D005290:Ferric Compounds; D026361:Reactive Nitrogen Species; D011994:Recombinant Proteins; D009569:Nitric Oxide; D003577:Cytochrome P-450 Enzyme System; C501935:cytochrome P-450 CYP51, Mycobacterium tuberculosis