Structure of mycobacterial NDH-2 bound to a 2-mercapto-quinazolinone inhibitor

The mycobacterial type II NADH dehydrogenase (NDH-2) is a promising drug target because of its central role in energy metabolism in Mycobacterium tuberculosis and other pathogens, and because it lacks a known mammalian homologue. However, the absence of structural information on how the enzyme binds inhibitors has made optimization of lead compounds challenging. We used electron cryomicroscopy (cryo-EM) to determine the structure of NDH-2 from Mycobacterium smegmatis , a fast-growing non-pathogenic model for respiration in M. tuberculosis , both alone and in complex with a 2-mercapto-quinazolinone inhibitor. The structure shows that active mycobacterial NDH-2 is dimeric, with the dimerization interface stabilized by an extended C-terminal α helix not found in NDH-2 from other bacterial genera. The arrangement of monomers in the dimer differs from the arrangement described for other prokaryotic NDH-2 dimers, instead resembling dimers formed by NDH-2 in eukaryotes. Density for the 2-mercapto-quinazolinone in the menaquinone-binding site shows that the inhibitor blocks menaquinone reduction through direct interaction with the flavin adenine dinucleotide cofactor. These results reveal structural elements of NDH-2 that could be used to design specific inhibitors of the mycobacterial enzyme.