The G‐protein FlhF has a role in polar flagellar placement and general stress response induction in <i>Pseudomonas putida</i>

Pandža, Suada; Baetens, Margot; Park, C. H.; Au, Timothy; Keyhan, M.; Матин, А.

doi:10.1046/j.1365-2958.2000.01859.x

Cited by 117 publications

(145 citation statements)

References 34 publications

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“…The recent structure of the SRP-FtsY complex, together with biochemical implications, suggest that the distal end of the hairpin-like SRP RNA may be involved in this activation 18 . The third SRP-GTPase FlhF, together with the MinD-type protein YlxH (also known as FlhG, FleN, motR or MinD2), is essential for the placement and assembly of flagella 19 in many polar and peritrichous flagellated bacteria [20][21][22][23][24] . FlhF is required for the targeting of the first flagellar protein, FliF, to the cell pole 25 by a mechanism that is so far poorly understood.…”

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confidence: 99%

Structural basis for the molecular evolution of SRP-GTPase activation by protein

Bange

Kümmerer

Grudnik

et al. 2011

Nat Struct Mol Biol

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SIMIBI-class (named after the signal recognition particle, MinD, BioD) nucleotide-binding proteins appeared early in evolution 1 and contain GTPases, as well as ATPases, involved in the correct localization of cellular constituents. The MinD ATPase, as the central part of the Min system, regulates the determination of the cell division site in all bacterial species 2 . SRP-GTPases form a subfamily of the SIMIBI class, with only three members: the signal sequence-binding protein Ffh (SRP54 in Eukarya and Archaea), the SRP receptor FtsY (SR in Eukarya) and FlhF, which is involved in flagella biosynthesis [3][4][5] . They share the conserved NG domain, which contains two major additions to the conserved fold of small G proteins. First, an --element (I-box) is inserted in the effector region; second, the N domain, comprising four -helices, is attached to the N terminus of the G domain. SRP (Ffh together with the SRP RNA) and FtsY constitute the universally conserved co-translational protein-targeting machinery 6,7 . When bound to GTP, Ffh and FtsY form, through interactions between their NG domains 8,9 , a heterodimeric complex that regulates the transfer of a ribosome-nascent chain complex to a vacant translocon in the membrane with a series of conformational rearrangements 10,11 . The two GTPases share a composite active site between their G domains in which GTP hydrolysis is reciprocally activated 12 . The SRP RNA [13][14][15] and membrane lipids 16,17 play fundamental roles in activating the Ffh-FtsY GTPases. The recent structure of the SRP-FtsY complex, together with biochemical implications, suggest that the distal end of the hairpin-like SRP RNA may be involved in this activation 18 . The third SRP-GTPase FlhF, together with the MinD-type protein YlxH (also known as FlhG, FleN, motR or MinD2), is essential for the placement and assembly of flagella 19 in many polar and peritrichous flagellated bacteria [20][21][22][23][24] . FlhF is required for the targeting of the first flagellar protein, FliF, to the cell pole 25 by a mechanism that is so far poorly understood. FlhF is associated with the membrane 25,26 and localizes at the cell pole 20 . The FlhF protein (Fig. 1a) contains an N-terminal B domain that seems to be involved in FliF targeting 25 ; it shares the NG domain fold with the other two members of the SRP-GTPase subfamily. FlhF forms a stable homodimer with GTP and a composite active site that is basically identical to the active site of the Ffh-FtsY heterodimer 5 . In both the homo-and heterodimer, the two nucleotides are bound in a head-to-tail manner, with the -phosphate of one nucleotide interacting with the 3 -OH of the ribose moiety of the other. However, for the homo-and heterodimers formed by the three SRP-GTPases, the molecular mechanism of activation is still unknown. We set out to understand the activation of SRPGTPases by studying FlhF. RESULTS The SRP-GTPase FlhF is activated by YlxHAs FlhF (Fig. 1) forms a stable homodimer, and reciprocal activation has not been observed 5 , we reasoned ...

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confidence: 99%

Structural basis for the molecular evolution of SRP-GTPase activation by protein

Bange

Kümmerer

Grudnik

et al. 2011

Nat Struct Mol Biol

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“…FlhF associates with the plasma membrane (7), where it localizes at the cell poles (6). Loss of FlhF induces a number of changes, from reduced transcription of flagellar genes to a random arrangement of flagella and even to absent or diminished motility (6,(8)(9)(10)(11)(12)(13). In yeast two-hybrid screens, FlhF was shown to dimerize (10).…”

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confidence: 99%

The crystal structure of the third signal-recognition particle GTPase FlhF reveals a homodimer with bound GTP

Bange

Petzold

Wild

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

101

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Flagella are well characterized as the organelles of locomotion and allow bacteria to react to environmental changes. The assembly of flagella is a multistep process and relies on a complex type III export machinery located in the cytoplasmic membrane. The FlhF protein is essential for the placement and assembly of polar flagella and has been classified as a signal-recognition particle (SRP)-type GTPase. SRP GTPases appeared early in evolution and form a unique subfamily within the guanine nucleotide binding proteins with only three members: the signal sequence-binding protein SRP54, the SRP receptor FtsY, and FlhF. We report the crystal structures of FlhF from Bacillus subtilis in complex with GTP and GMPPNP. FlhF shares SRP GTPase-specific features such as the presence of an N-terminal ␣-helical domain and the I-box insertion. It forms a symmetric homodimer sequestering a composite active site that contains two head-to-tail arranged nucleotides similar to the heterodimeric SRP-targeting complex. However, significant differences to the GTPases of SRP and the SRP receptor include the formation of a stable homodimer with GTP as well as severe modifications and even the absence of motifs involved in regulation of the other two SRP GTPases. Our results provide insights into SRP GTPases and their roles in two fundamentally different protein-targeting routes that both rely on efficient protein delivery to a secretion channel.flagellum ͉ protein translocation ͉ x-ray structure analysis ͉ protein targeting

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“…PflI appears to function differently from FlhF, which has been identified for its role in polar flagellar placement in several species (38,41). How FlhF affects flagellum positioning is not well understood, but the sequence similarity between FlhF and GTP-binding SRP proteins suggests a possible role in translocation of early flagellar proteins.…”

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confidence: 99%

“…It has been shown that in several polarly flagellated species of gammaproteobacteria, the flhF gene product is necessary for restricting the flagellum to a polar position (38,41). flhF encodes a GTP-binding protein homologous to components of the signal recognition particle (SRP) pathway; in its absence, Pseudomonas putida and Pseudomonas aeruginosa have reduced motility and randomly placed flagella on the cell surface (25,38,41).…”

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confidence: 99%

“…flhF encodes a GTP-binding protein homologous to components of the signal recognition particle (SRP) pathway; in its absence, Pseudomonas putida and Pseudomonas aeruginosa have reduced motility and randomly placed flagella on the cell surface (25,38,41). Surprisingly, flhF homologs are also found in species with multiple, randomly placed flagella, indicating that FlhF is not a defining component of polarly flagellated bacteria.…”

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PflI, a Protein Involved in Flagellar Positioning in Caulobacter crescentus

Obuchowski

Jacobs‐Wagner

2008

J Bacteriol

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The bacterial flagellum is important for motility and adaptation to environmental niches. The sequence of events required for the synthesis of the flagellar apparatus has been extensively studied, yet the events that dictate where the flagellum is placed at the onset of flagellar biosynthesis remain largely unknown. We addressed this question for alphaproteobacteria by using the polarly flagellated alphaproteobacterium Caulobacter crescentus as an experimental model system. To identify candidates for a role in flagellar placement, we searched all available alphaproteobacterial genomes for genes of unknown function that cluster with early flagellar genes and that are present in polarly flagellated alphaproteobacteria while being absent in alphaproteobacteria with other flagellation patterns. From this in silico screen, we identified pflI. Loss of PflI function in C. crescentus results in an abnormally high frequency of cells with a randomly placed flagellum, while other aspects of cell polarization remain normal. In a wild-type background, a fusion of green fluorescent protein (GFP) and PflI localizes to the pole where the flagellum develops. This polar localization is independent of the flagellar protein FliF, whose oligomerization into the MS ring is thought to define the site of flagellar synthesis, suggesting that PflI acts before or independently of this event. Overproduction of PflI-GFP often leads to ectopic localization at the wrong, stalked pole. This is accompanied by a high frequency of flagellum formation at this ectopic site, suggesting that the location of PflI is a sufficient marker for a site for flagellar assembly.Flagellar arrangement has long been used as a defining characteristic in bacterial taxonomy. Bacteria can have flagella emerging from a specific site, such as the cell pole, or from seemingly random positions along the cell body (as in peritrichously flagellated bacteria). Differences in flagellar distribution can be important for how various species adapt to specific environments. While the biosynthesis of the flagellum and the temporal regulation of its assembly have been extensively studied (5, 32), we know little about what controls the cellular position of flagellar assembly. The distribution of flagella in peritrichous bacteria may be random and stochastically driven, but spatial mechanisms must exist in polarly flagellated bacteria to ensure that flagellar assembly takes place at the right location.It has been shown that in several polarly flagellated species of gammaproteobacteria, the flhF gene product is necessary for restricting the flagellum to a polar position (38, 41). flhF encodes a GTP-binding protein homologous to components of the signal recognition particle (SRP) pathway; in its absence, Pseudomonas putida and Pseudomonas aeruginosa have reduced motility and randomly placed flagella on the cell surface (25,38,41). Surprisingly, flhF homologs are also found in species with multiple, randomly placed flagella, indicating that FlhF is not a defining component of polarly f...

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The G‐protein FlhF has a role in polar flagellar placement and general stress response induction in Pseudomonas putida

Cited by 117 publications

References 34 publications

Structural basis for the molecular evolution of SRP-GTPase activation by protein

Structural basis for the molecular evolution of SRP-GTPase activation by protein

The crystal structure of the third signal-recognition particle GTPase FlhF reveals a homodimer with bound GTP

PflI, a Protein Involved in Flagellar Positioning in Caulobacter crescentus

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