Novel targets and inhibitors of the Mycobacterium tuberculosis cytochrome bd oxidase to foster anti-tuberculosis drug discovery.

INTRODUCTION: Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), is the most devastating bacterial disease. Multidrug-resistant Mtb strains are spreading worldwide, underscoring the need for new anti-TB targets and inhibitors. The respiratory chain complexes, including the cytochrome bd oxidase (cyt-bd), have been identified as an attractive target for drug development. Recent novel structural and mechanistic insight as well as inhibitors of Mtb's cyt-bd brought this enzyme into the focus. AREAS COVERED: In this review, the authors describe conditions that stimulate the biogenesis of Mtb cyt-bd, its structural-, mechanistic-, and substrate-binding traits. They discuss the present Mtb cyt-bd inhibitors, novel targets within the enzyme and structure activity relationship features that are required for mycobacterial cyt-bd inhibition and augment their understanding on improving the potency of cyt-bd inhibitors. EXPERT OPINION: A deeper structure-mechanistic understanding of Mtb's cyt-bd is a prerequisite for in silico efforts to: (i) identify pathogen specific targets for the design of novel nontoxic hit molecules, forming the platform for the development of new leads, (ii) design mechanism of action studies, (iii) perform medicinal chemistry of existing inhibitors to improve their potency and pharmacokinetic/-dynamic properties. Phase studies with such optimized cyt-bd inhibitors in combination with anti-TB compounds targeting the oxidative phosphorylation pathway is recommended.

Cryo-Electron Microscopy Structure of the Mycobacterium tuberculosis Cytochrome bcc:aa3 Supercomplex and a Novel Inhibitor Targeting Subunit Cytochrome cI.

The mycobacterial cytochrome bcc:aa3 complex deserves the name "supercomplex" since it combines three cytochrome oxidases-cytochrome bc, cytochrome c, and cytochrome aa3-into one supramolecular machine and performs electron transfer for the reduction of oxygen to water and proton transport to generate the proton motive force for ATP synthesis. Thus, the bcc:aa3 complex represents a valid drug target for Mycobacterium tuberculosis infections. The production and purification of an entire M. tuberculosis cytochrome bcc:aa3 are fundamental for biochemical and structural characterization of this supercomplex, paving the way for new inhibitor targets and molecules. Here, we produced and purified the entire and active M. tuberculosis cyt-bcc:aa3 oxidase, as demonstrated by the different heme spectra and an oxygen consumption assay. The resolved M. tuberculosis cyt-bcc:aa3 cryo-electron microscopy structure reveals a dimer with its functional domains involved in electron, proton, oxygen transfer, and oxygen reduction. The structure shows the two cytochrome cIcII head domains of the dimer, the counterpart of the soluble mitochondrial cytochrome c, in a so-called "closed state," in which electrons are translocated from the bcc to the aa3 domain. The structural and mechanistic insights provided the basis for a virtual screening campaign that identified a potent M. tuberculosis cyt-bcc:aa3 inhibitor, cytMycc1. cytMycc1 targets the mycobacterium-specific α3-helix of cytochrome cI and interferes with oxygen consumption by interrupting electron translocation via the cIcII head. The successful identification of a new cyt-bcc:aa3 inhibitor demonstrates the potential of a structure-mechanism-based approach for novel compound development.