Co-Folding of a FliF-FliG Split Domain Forms the Basis of the MS:C Ring Interface within the Bacterial Flagellar Motor

Lynch, Michael J.; Levenson, R.; Kim, Eun A; Sircar, Ria; Blair, David F.; Dahlquist, Frederick W.; Crane, Brian R.

doi:10.1016/j.str.2016.12.006

Cited by 66 publications

(102 citation statements)

References 39 publications

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Section: Downloaded Fromsupporting

confidence: 90%

“…The interruption of the two salt bridges, R529 F l i F :E87 F l i G and K553 F l i F :D42 F l i G , did not have any effect on the heterodimer formation, suggesting that the hydrophobic contacts were the dominant contributors to the stable interaction of FliF and FliG. These results agree with earlier mutagenesis studies in E. coli and Salmonella and with the structure of T. maritima 6,[10][11][12].Flagellar formation and full motility required hydrophobic contacts between FliF and FliG Because our structure represented the first atomic model of the FliF-FliG complex from mesophilic bacteria, we completed an in vivo complementation study in an E. coli system to confirm the importance of the hydrophobic contacts between FliF C and FliG N in flagellar motor assembly and function. Various FliG constructs were created: F59D, F66D and F59D/F66D.…”

supporting

confidence: 90%

“…Many biochemical studies have shown that the junction of the MS and C rings arises from the 1:1 stoichiometric interaction between the C-terminal 40-residue long cytoplasmic region of FliF (FliF C ) and the N-terminal domain of FliG (FliG N ; [4][5][6][7][8][9][10][11]. The characterization of spontaneous FliF-FliG deletionfusion mutants in Salmonella and other mutagenesis studies have further shown that gene interruption of FliF C and FliG N cause a defect in flagellation and motility (4)(5)(6)12), which strongly suggests that the optimal function of flagella requires the interaction and conformational freedom of FliF C and FliG N .…”

mentioning

confidence: 99%

“…However, the molecular details of the assembly of FliG and FliF in the inner ring are also poorly understood. Currently, the information about FliG N is only available from the homologs of the thermophile Aquifex aeolicus and Thermotoga maritima (10,13), and the corresponding structure in human pathogen-related mesophilic bacteria remains unresolved. Notably, the structural model of the FliG N ring generated from the crystal structure of FliFFliG from T. maritima did not reconcile with the E. coli model derived from the biochemical studies (10,11).…”

mentioning

confidence: 99%

“…Currently, the information about FliG N is only available from the homologs of the thermophile Aquifex aeolicus and Thermotoga maritima (10,13), and the corresponding structure in human pathogen-related mesophilic bacteria remains unresolved. Notably, the structural model of the FliG N ring generated from the crystal structure of FliFFliG from T. maritima did not reconcile with the E. coli model derived from the biochemical studies (10,11). During our previous efforts to solve the structure of FliG M C from Helicobacter pylori, a human pathogenic bacterium related to gastric cancer, we noticed that while the middle and C-terminal domains of H. pylori, FliG shared a relatively high sequence identity with their thermophilic counterparts, the N-terminal domain of FliGN was less conserved (about 18% and 30% sequence identity with that from A. aeolicus and T. maritima, respectively).…”

mentioning

confidence: 99%

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Crystal structure of the FliF–FliG complex from Helicobacter pylori yields insight into the assembly of the motor MS–C ring in the bacterial flagellum

Xue

Lam

Zhang

et al. 2018

Journal of Biological Chemistry

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The bacterial flagellar motor is a self-assembling supramolecular nanodevice.Its spontaneous biosynthesis is initiated by the insertion of the MS ring protein FliF into the inner membrane, followed by attachment of the switch protein FliG. Assembly of this multiprotein complex is tightly regulated to avoid nonspecific aggregation, but the molecular mechanisms governing flagellar assembly are unclear. Here, we present the crystal structure of the cytoplasmic domain of FliF complexed with the Nterminal domain of FliG (FliF C -FliG N ) from the bacterium Helicobacter pylori. Within this complex, FliF C interacted with FliG N through extensive hydrophobic contacts similar to those observed in the FliF C -FliG N structure from the thermophile Thermotoga maritima, indicating conservation of the FliF C -FliG N interaction across bacterial species. Analysis of the crystal lattice revealed that the heterodimeric complex packs as a linear superhelix via stacking of the armadillo-repeat-like motifs (ARM) of FliG N . Notably, this linear helix was similar to that observed for the assembly of the FliG middle domain. We validated the in vivo relevance of the FliG N stacking by complementation studies in Escherichia coli. Furthermore, structural comparison with apo FliG from the thermophile Aquifex aeolicus indicated that FliF regulates the conformational transition of FliG and exposes the complementary ARM-like motifs of FliG N , containing conserved hydrophobic residues. FliF apparently both provides a template for FliG polymerization and spatiotemporally controls subunit interactions within FliG. Our findings reveal that a small protein fold can serve as a versatile building block to assemble into a multiprotein machinery of distinct shapes for specific functions.The bacterial flagellar motor is a self-assembled reversible rotary nanodevice. This dynamic rotary motor of about 40 nm in diameter is composed of rings of protein oligomers: the L ring (outer membrane), P ring (peptidoglycan layer), MS ring (inner membrane) and C ring (cytoplasm) (1,2). The L and P rings are believed to act as a bushing through which a central rotating rod can pass. The MS and C rings contribute to the rotor part of the flagellar motor. Torque is generated by a membrane-bound stator, which converts electrochemical potential into mechanical force that acts on the rotor. The synthesis of the flagellum begins at the rotor, which is composed of the MS and C ring protein subassemblies ( Figure 1). Specifically, the assembly of the MS ring protein FliF prompts the incorporation of the switch protein FliG and subsequently the proteins FliM and FliN to form the cytoplasmic motor switch complex. The MS ring plays an important role as it interacts both with the rod where the flagellum is anchored and with the C ring where torque Crystal structure of FliF-FliG complex from H. pylori generation and rotation switching take place.In Salmonella typhimurium and Escherichia coli, there are about 26 copies of FliF in the MS ring and 26 copies of FliG, 34 c...

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Section: Downloaded Fromsupporting

confidence: 90%

supporting

confidence: 90%