Crystal structures of F420-dependent glucose-6-phosphate dehydrogenase FGD1 involved in the activation of the anti-tuberculosis drug candidate PA-824 reveal the basis of coenzyme and substrate binding

The modified flavin coenzyme F-420 is found in a restricted number of microorganisms. It is widely distributed in mycobacteria, however, where it is important in energy metabolism, and in Mycobacterium tuberculosis (Mtb) is implicated in redox processes related to non-replicating persistence. In Mtb, the F-420-dependent glucose-6-phosphate dehydrogenase FGD1 provides reduced F-420 for the in vivo activation of the nitroimidazopyran prodrug PA-824, currently being developed for anti-tuberculosis therapy against both replicating and persistent bacteria. The structure of M. tuberculosis FGD1 has been determined by x-ray crystallography both in its apo state and in complex with F-420 and citrate at resolutions of 1.90 and 1.95 angstrom, respectively. The structure reveals a highly specific F420 binding mode, which is shared with several other F-420-dependent enzymes. Citrate occupies the substrate binding pocket adjacent to F-420 and is shown to be a competitive inhibitor (IC50 43 mu M). Modeling of the binding of the glucose 6-phosphate (G6P) substrate identifies a positively charged phosphate binding pocket and shows that G6P, like citrate, packs against the isoalloxazine moiety of F-420 and helps promote a butterfly bend conformation that facilitates F-420 reduction and catalysis.