Structural insights into the functional versatility of an FHA domain protein in mycobacterial signaling

• DOI: 10.1126/scisignal.aav9504
• Publication Date: 2019-05-07T23:15:20Z
• Authors: Tristan Wagner; Gwénaëlle André-L...; Valérie Hindie; Nathalie Barilone; María-Natalia Lisa; Sylviane Hoos; Bertrand Raynal; Brigitte Vulliez-...; Helen M. O’Hare; Marco Bellinzoni; Pedro M. Alzari
• Keywords: Models, Molecular; 0301 basic medicine; 570; phosphomimetics; Carboxy-Lyases; [SDV]Life Sciences [q-bio]; Protein Serine-Threonine Kinases; Crystallography, X-Ray; PknB; 03 medical and health sciences; Bacterial Proteins; https://purl.org/becyt/ford/1.6; Amino Acid Sequence; Phosphorylation; https://purl.org/becyt/ford/1; phosphothreonine; GarA; Antigens, Bacterial; Binding Sites; Forkhead Transcription Factors; Mycobacterium tuberculosis; Forkhead-associated; Protein Structure, Tertiary; Mutation; Protein Binding; Signal Transduction

Forkhead-associated (FHA) domains are modules that bind to phosphothreonine (pThr) residues in signaling cascades. The FHA-containing mycobacterial protein GarA is a central element of phosphorylation-dependent pathway redirects metabolic flux response amino acid starvation or cell growth requirements. acts as ON/OFF molecular switch. In its nonphosphorylated ON state, the FHA domain engages phosphorylation-independent interactions with various enzymes orchestrate nitrogen flow, such 2-oxoglutarate decarboxylase (KGD). However, phosphorylation at N-terminal region by kinase PknB PknG triggers autoinhibition through intramolecular association domain, thus blocking all downstream interactions. To investigate these different binding modes, we solved crystal structures upstream (phosphorylation-dependent) complex PknB-GarA and (phosphorylation-independent) GarA-KGD. Our results show phosphorylated activation loop serves docking site recruit canonical FHA-pThr same FHA-binding pocket targets an allosteric on KGD, where key recognition phosphomimetic aspartate. Further enzymatic mutagenesis studies revealed acted dynamic inhibitor KGD preventing crucial motions necessary for catalysis. provide evidence physiological phosphomimetics, supporting numerous using approaches, illustrate how evolution can shape single specifically interact multiple partners.