Structural Insights into the Interaction between the Bacterial Flagellar Motor Proteins FliF and FliG

• Published in: Biochemistry (Easton) 2012-06, Vol.51 (25), p.5052-5060
• Main Authors: Levenson, Robert, Zhou, Hongjun, Dahlquist, Frederick W
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Bacteria; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Caulobacter crescentus - chemistry; Flagella - chemistry; Flagella - genetics; Flagella - metabolism; Membrane Proteins - chemistry; Membrane Proteins - genetics; Membrane Proteins - metabolism; Molecular Motor Proteins - chemistry; Molecular Motor Proteins - genetics; Molecular Motor Proteins - metabolism; Molecular Sequence Data; Protein Binding; Thermotoga maritima; Thermotoga maritima - chemistry
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi3004582

The binding of the soluble cytoplasmic protein FliG to the transmembrane protein FliF is one of the first interactions in the assembly of the bacterial flagellum. Once established, this interaction is integral in keeping the flagellar cytoplasmic ring, responsible for both transmission of torque and control of the rotational direction of the flagellum, anchored to the central transmembrane ring on which the flagellum is assembled. Here we isolate and characterize the interaction between the N-terminal domain of Thermotoga maritima FliG (FliGN) and peptides corresponding to the conserved C-terminal portion of T. maritima FliF. Using nuclear magnetic resonance (NMR) and other techniques, we show that the last ∼40 amino acids of FliF (FliFC) interact strongly (upper bound K d in the low nanomolar range) with FliGN. The formation of this complex causes extensive conformational changes in FliGN. We find that T. maritima FliGN is homodimeric in the absence of the FliFC peptide but forms a heterodimeric complex with the peptide, and we show that this same change in oligomeric state occurs in full-length T. maritima FliG, as well. We relate previously observed phenotypic effects of FliFC mutations to our direct observation of binding. Lastly, on the basis of NMR data, we propose that the primary interaction site for FliFC is located on a conserved hydrophobic patch centered along helix 1 of FliGN. These results provide new detailed information about the bacterial flagellar motor and support efforts to understand the cytoplasmic ring’s precise molecular structure and mechanism of rotational switching.