The Dimer-of-Trimers Assembly Prevents Catalysis at the Transferase Site of Prokaryotic FAD Synthase.
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ABSTRACT: Flavin mononucleotide (FMN) and flavin-adenine dinucleotide (FAD) are essential flavoprotein cofactors. A riboflavin kinase (RFK) activity catalyzes riboflavin phosphorylation to FMN, which can then be transformed into FAD by an FMN:adenylyltransferase (FMNAT) activity. Two enzymes are responsible for each one of these activities in eukaryotes, whereas prokaryotes have a single bifunctional enzyme, FAD synthase (FADS). FADS folds in two independent modules: the C-terminal with RFK activity and the N-terminal with FMNAT activity. Differences in structure and chemistry for the FMNAT catalysis among prokaryotic and eukaryotic enzymes pointed to the FMNAT activity of prokaryotic FADS as a potential antimicrobial target, making the structural model of the bacterial FMNAT module in complex with substrates relevant to understand the FADS catalytic mechanism and to the discovery of antimicrobial drugs. However, such a crystallographic complex remains elusive. Here, we have used molecular docking and molecular dynamics simulations to generate energetically stable interactions of the FMNAT module of FADS from Corynebacterium ammoniagenes with ATP/Mg2+ and FMN in both the monomeric and dimer-of-trimers assemblies reported for this protein. For the monomer, we have identified the residues that accommodate the reactive phosphates in a conformation compatible with catalysis. Interestingly, for the dimer-of-trimers conformation, we have found that the RFK module negatively influences FMN binding at the interacting FMNAT module. These results agree with calorimetric data of purified samples containing nearly 100% monomer or nearly 100% dimer-of-trimers, indicating that FMN binds to the monomer but not to the dimer-of-trimers. Such observations support regulation of flavin homeostasis by quaternary C. ammoniagenes FADS assemblies.
SUBMITTER: Lans I
PROVIDER: S-EPMC6139884 | biostudies-literature | 2018 Sep
REPOSITORIES: biostudies-literature
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