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1.
Genetic and molecular characterization of the polar flagellum of Vibrio parahaemolyticus. 总被引:12,自引:5,他引:7
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L L McCarter 《Journal of bacteriology》1995,177(6):1595-1609
Vibrio parahaemolyticus possesses two alternate flagellar systems adapted for movement under different circumstances. A single polar flagellum propels the bacterium in liquid (swimming), while multiple lateral flagella move the bacterium over surfaces (swarming). Energy to rotate the polar flagellum is derived from the sodium membrane potential, whereas lateral flagella are powered by the proton motive force. Lateral flagella are arranged peritrichously, and the unsheathed filaments are polymerized from a single flagellin. The polar flagellum is synthesized constitutively, but lateral flagella are produced only under conditions in which the polar flagellum is not functional, e.g., on surfaces. This work initiates characterization of the sheathed, polar flagellum. Four genes encoding flagellins were cloned and found to map in two loci. These genes, as well as three genes encoding proteins resembling HAPs (hook-associated proteins), were sequenced. A potential consensus polar flagellar promoter was identified by using upstream sequences from seven polar genes. It resembled the enterobacterial sigma 28 consensus promoter. Three of the four flagellin genes were expressed in Escherichia coli, and expression was dependent on the product of the fliA gene encoding sigma 28. The fourth flagellin gene may be different regulated. It was not expressed in E. coli, and inspection of upstream sequence revealed a potential sigma 54 consensus promoter. Mutants with single and multiple defects in flagellin genes were constructed in order to determine assembly rules for filament polymerization. HAP mutants displayed new phenotypes, which were different from those of Salmonella typhimurium and most probably were the result of the filament being sheathed. 相似文献
2.
Assembly of the long helical filament of the bacterial flagellum requires polymerisation of ca 20,000 flagellin (FliC) monomeric subunits into the growing structure extending from the cell surface. Here, we show that export of Salmonella flagellin is facilitated specifically by a cytosolic protein, FliS, and that FliS binds to the FliC C-terminal helical domain, which contributes to stabilisation of flagellin subunit interactions during polymerisation. Stable complexes of FliS with flagellin were assembled efficiently in vitro, apparently by FliS homodimers binding to FliC monomers. The data suggest that FliS acts as a substrate-specific chaperone, preventing premature interaction of newly synthesised flagellin subunits in the cytosol. Compatible with this view, FliS was able to prevent in vitro polymerisation of FliC into filaments. 相似文献
3.
4.
G W Ordal 《Journal of bacteriology》1976,125(3):1005-1012
Bacillus subtilis, like Escherichia coli and Salmonella typhimurium, carries out chemotaxis by modulating the relative frequency of smooth swimming and tumbling. Like these enteric bacteria, methionine auxotrophs starved for methionine show an abnormally long-period of smooth swimming after addition of attractant. This "hypersensitive" state requires an hour of starvation for its genesis, which can be hastened by including alanine, a strong attractant, in starvation medium. Susceptibility to repellent, which causes transient tumbling when added, if anything, increases slightly by starvation for methionine. The results are interpreted by postulating the existence of a methionine-derived structure that hastens recovery of attractant-stimulated bacteria back to normal. 相似文献
5.
Furukawa Y Imada K Vonderviszt F Matsunami H Sano K Kutsukake K Namba K 《Journal of molecular biology》2002,318(3):889-900
The axial structure of the bacterial flagellum is composed of many different proteins, such as hook protein and flagellin, and each protein forms a short or long axial segment one after another in a well-defined order along the axis. Under physiological conditions, most of these proteins are stable in the monomeric state in solution, and spontaneous polymerization appears to be suppressed, as demonstrated clearly for flagellin, probably to avoid undesirable self-assembly in the cytoplasmic space. However, no systematic studies of the possible associations between monomeric axial proteins in solution have been carried out. We therefore studied self and cross-association between hook protein, flagellin and three hook-associated proteins, HAP1, HAP2 and HAP3, in all possible pairs, by gel-filtration and analytical centrifugation, and found interactions in the following two cases only. Flagellin facilitated HAP3 aggregation into beta-amyloid-like filaments, but without stable binding between the two. Addition of HAP3 to HAP2 resulted in disassembly of preformed HAP2 decamers and formation of stable HAP2-HAP3 heterodimers. HAP2 missing either of its disordered terminal regions did not form the heterodimer, whereas HAP3 missing either of its disordered terminal regions showed stable heterodimer formation. This polarity in the heterodimer interactions suggests that the interactions between HAP2 and HAP3 in solution are basically the same as those in the flagellar axial structure. We discuss these results in relation to the assembly mechanism of the flagellum. 相似文献
6.
C.R. Calldine 《Journal of molecular biology》1978,118(4):457-479
The flagella of Salmonella and other bacteria are constructed from molecules of the protein flagellin in a way which permits relatively easy transition between members of a family of distinct stable left and right-handed helical waveforms. Changes of waveform, particularly between “normal” (left-handed) and “curly” (right-handed) play an important role in the switch from smooth swimming to tumbling in chemotaxis. This paper establishes some mechanical properties of model flagella built from bi-stable subunits, which in turn clarifies the mechanics of the changes of waveform which occur, in a viscous fluid environment, at various points in the swimming cycle.Available data on the joining of different helical waveforms in a single filament, supplemented by information on the way in which helical filaments flatten down in preparation for electron microscopy, are well-fitted by the mechanical behaviour of an assembly of mechanical subunits having some simple distinctive design features. The same arrangement makes possible an explanation for the formation of flagellar-like but straight polymers from Salmonella flagellin in the presence of high concentrations of NaCl. 相似文献
7.
Pyatibratov MG Beznosov SN Rachel R Tiktopulo EI Surin AK Syutkin AS Fedorov OV 《Canadian journal of microbiology》2008,54(10):835-844
Many Archaea use rotation of helical flagellar filaments for swimming motility. We isolated and characterized the flagellar filaments of Haloarcula marismortui, an archaeal species previously considered to be nonmotile. Two Haloarcula marismortui phenotypes were discriminated--their filaments are composed predominantly of either FlaB or FlaA2 flagellin, and the corresponding genes are located on different replicons. FlaB and FlaA2 filaments differ in antigenicity and thermostability. FlaA2 filaments are distinctly thicker (20-22 nm) than FlaB filaments (16-18 nm). The observed filaments are nearly twice as thick as those of other characterized euryarchaeal filaments. The results suggest that the helicity of Haloarcula marismortui filaments is provided by a mechanism different from that in the related haloarchaeon Halobacterium salinarum, where 2 different flagellin molecules present in comparable quantities are required to form a helical filament. 相似文献
8.
G Kuwajima 《Journal of bacteriology》1988,170(1):485-488
Escherichia coli K-12 mutants with altered flagellum antigenicity were isolated by introducing random deletions into the flagellin gene. The deletions were identified in the central region of the gene. It is suggested that this region corresponds to the flagellin domain molecule which affects flagellum antigenicity. 相似文献
9.
Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of flagella dissociated from strains of Campylobacter coli and Campylobacter jejuni belonging to the heat-labile serogroup LIO 8 showed that some strains were capable of producing flagellin subunits of two different molecular weights (MrS), 59,500 and 61,500. Immunoelectron microscopy of cultures of the type strain of this serogroup, C. coli VC167, showed the presence of two flagellum filaments of different antigenic specificity. Epitopes on the surface of one of these flagella bound antibodies in LIO 8 typing antiserum, and Western blotting (immunoblotting) and immunoprecipitation showed that the flagellum was composed of flagellin of Mr 61,500. The other flagellum antigenic type did not bind LIO 8 antibodies but did possess serospecific epitopes which bound a second polyclonal antiserum, LAH2. This second antigenic flagellum type was composed of the Mr 59,500 flagellin. Cells producing either of the flagellum antigenic types serotyped as LIO 8, indicating that flagella composed of the Mr 61,500 flagellin do not carry the serological determinants for this serogroup. The ability of C. coli VC167 to produce these flagella of different subunit MrS was shown to represent a bidirectional antigenic variation. When measured in culture medium, the phase 1-to-phase 2 transition occurred at a rate of approximately 2.0 x 10(-5) per cell per generation, and the phase 2-to-phase 1 transition occurred at a rate of 1.2 x 10(-6) per cell per generation. 相似文献
10.
Pausing of flagellar rotation is a component of bacterial motility and chemotaxis. 总被引:13,自引:4,他引:9
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When bacterial cells are tethered to glass by their flagella, many of them spin. On the basis of experiments with tethered cells it has generally been thought that the motor which drives the flagellum is a two-state device, existing in either a counterclockwise or a clockwise state. Here we show that a third state of the motor is that of pausing, the duration and frequency of which are affected by chemotactic stimuli. We have recorded on video tape the rotation of tethered Escherichia coli and Salmonella typhimurium cells and analyzed the recordings frame by frame and in slow motion. Most wild-type cells paused intermittently. The addition of repellents caused an increase in the frequency and duration of the pauses. The addition of attractants sharply reduced the number of pauses. A chemotaxis mutant which lacks a large part of the chemotaxis machinery owing to a deletion of the genes from cheA to cheZ did not pause at all and did not respond to repellents by pausing. A tumbly mutant of S. typhimurium responded to repellents by smooth swimming and to attractants by tumbling. When tethered, these cells exhibited a normal rotational response but an inverse pausing response to chemotactic stimuli: the frequency of pauses decreased in response to repellents and increased in response to attractants. It is suggested that (i) pausing is an integral part of bacterial motility and chemotaxis, (ii) pausing is independent of the direction of flagellar rotation, and (iii) pausing may be one of the causes of tumbling. 相似文献
11.
Flagellar motility and structure in the hyperthermoacidophilic archaeon Sulfolobus solfataricus
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Szabó Z Sani M Groeneveld M Zolghadr B Schelert J Albers SV Blum P Boekema EJ Driessen AJ 《Journal of bacteriology》2007,189(11):4305-4309
Flagellation in archaea is widespread and is involved in swimming motility. Here, we demonstrate that the structural flagellin gene from the crenarchaeaon Sulfolobus solfataricus is highly expressed in stationary-phase-grown cells and under unfavorable nutritional conditions. A mutant in a flagellar auxiliary gene, flaJ, was found to be nonmotile. Electron microscopic imaging of the flagellum indicates that the filaments are composed of right-handed helices. 相似文献
12.
Flores H Lobaton E Méndez-Diez S Tlupova S Cortez R 《Bulletin of mathematical biology》2005,67(1):137-168
Certain bacteria, such as Escherichia coli (E. coli) and Salmonella typhimurium (S. typhimurium), use multiple flagella often concentrated at one end of their bodies to induce locomotion. Each flagellum is formed in a
left-handed helix and has a motor at the base that rotates the flagellum in a corkscrew motion.We present a computational
model of the flagellar motion and their hydrodynamic interaction. The model is based on the equations of Stokes flow to describe
the fluid motion. The elasticity of the flagella is modeled with a network of elastic springs while the motor is represented
by a torque at the base of each flagellum. The fluid velocity due to the forces is described by regularized Stokeslets and
the velocity due to the torques by the associated regularized rotlets. Their expressions are derived. The model is used to
analyze the swimming motion of a single flagellum and of a group of three flagella in close proximity to one another. When
all flagellar motors rotate counterclockwise, the hydrodynamic interaction can lead to bundling. We present an analysis of
the flow surrounding the flagella. When at least one of the motors changes its direction of rotation, the same initial conditions
lead to a tumbling behavior characterized by the separation of the flagella, changes in their orientation, and no net swimming
motion. The analysis of the flow provides some intuition for these processes. 相似文献
13.
Rhodobacter sphaeroides can swim toward a wide range of attractants (a process known as taxis), propelled by a single rotating flagellum. The reversals of motor direction that cause tumbles in Eschericia coli taxis are replaced by brief motor stops, and taxis is controlled by a complex sensory system with multiple homologues of the E. coli sensory proteins. We tethered photosynthetically grown cells of R. sphaeroides by their flagella and measured the response of the flagellar motor to changes in light intensity. The unstimulated bias (probability of not being stopped) was significantly larger than the bias of tethered E. coli but similar to the probability of not tumbling in swimming E. coli. Otherwise, the step and impulse responses were the same as those of tethered E. coli to chemical attractants. This indicates that the single motor and multiple sensory signaling pathways in R. sphaeroides generate the same swimming response as several motors and a single pathway in E. coli, and that the response of the single motor is directly observable in the swimming pattern. Photo-responses were larger in the presence of cyanide or the uncoupler carbonyl cyanide 4-trifluoromethoxyphenylhydrazone (FCCP), consistent with the photo-response being detected via changes in the rate of electron transport. 相似文献
14.
The flagellar filament of Rhodobacter sphaeroides: pH-induced polymorphic transitions and analysis of the fliC gene
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Flagellar motility in Rhodobacter sphaeroides is notably different from that in other bacteria. R. sphaeroides moves in a series of runs and stops produced by the intermittent rotation of the flagellar motor. R. sphaeroides has a single, plain filament whose conformation changes according to flagellar motor activity. Conformations adopted during swimming include coiled, helical, and apparently straight forms. This range of morphological transitions is larger than that in other bacteria, where filaments alternate between left- and right-handed helical forms. The polymorphic ability of isolated R. sphaeroides filaments was tested in vitro by varying pH and ionic strength. The isolated filaments could form open-coiled, straight, normal, or curly conformations. The range of transitions made by the R. sphaeroides filament differs from that reported for Salmonella enterica serovar Typhimurium. The sequence of the R. sphaeroides fliC gene, which encodes the flagellin protein, was determined. The gene appears to be controlled by a sigma(28)-dependent promoter. It encodes a predicted peptide of 493 amino acids. Serovar Typhimurium mutants with altered polymorphic ability usually have amino acid changes at the terminal portions of flagellin or a deletion in the central region. There are no obvious major differences in the central regions to explain the difference in polymorphic ability. In serovar Typhimurium filaments, the termini of flagellin monomers have a coiled-coil conformation. The termini of R. sphaeroides flagellin are predicted to have a lower probability of coiled coils than are those of serovar Typhimurium flagellin. This may be one reason for the differences in polymorphic ability between the two filaments. 相似文献
15.
Identification of a sequence in human toll-like receptor 5 required for the binding of Gram-negative flagellin 总被引:8,自引:0,他引:8
Flagellins from Gram-negative bacteria activate inflammatory cells by a toll-like receptor 5 (TLR5)-dependent signaling pathway. We have examined the interaction between flagellin and TLR5 using an in vitro binding assay. Purified recombinant His-tagged flagellin from Salmonella enteritidis bound to TLR5 in detergent lysates from COS-1 cells transiently transfected with a human TLR5 expression plasmid. Flagellins from Salmonella typhimurium and Escherichia coli also bound to TLR5. The specificity of this interaction was demonstrated by its concentration dependence and lack of TLR5 binding to a biologically inactive form of flagellin or to a His-tagged non-flagellar protein. Flagellin bound to the extracellular domain of TLR5 expressed on the surface of COS-1 cells and to a soluble, monomeric form of the extracellular domain (amino acids 1-636). Although a TLR5 extracellular domain containing amino acids 1-407 retained flagellin binding activity, binding was not evident with a TLR5 peptide encoding residues 1-386. Conversely, a peptide containing amino acid residues 386-636 retained flagellin binding. Thus it is likely that amino acids 386-407 is a binding site for flagellin. This sequence contains a putative leucine-rich repeat. These results support the conclusion that flagellin signaling via TLR5 involves a direct interaction between flagellin and a leucine-rich region in TLR5. We also show that the NH2-terminal 358 amino acids of TLR5 play an important role in its signaling activity. Our results provide, for the first time, a molecular basis for the agonist specificity of a TLR. 相似文献
16.
Amino acids responsible for flagellar shape are distributed in terminal regions of flagellin 总被引:18,自引:0,他引:18
The shape of the flagellar filaments of the bacterium Salmonella typhimurium under ordinary conditions is a left-handed helix. In addition to the normal wild-type filament, non-helical (i.e. straight), right-handed helical (early), or circular (semi-coiled and coiled) filaments and filament with small amplitude (fl-type) have been found in mutants or in filaments reconstituted in vitro. We analysed wild-type flagellin and flagellins from 17 flagellar-shape mutants (6 with straight filaments, 6 with curly filaments, 4 with coiled filaments and 1 with fl-type filament) by amino acid sequencing to identify the mutational sites. All mutant flagellins except that of the fl-type filament had single mutations; the fl-type flagellin had two mutations in the molecule. The sites of these mutations were localized in alpha-helical segments of the terminal regions of flagellin. A possible mechanism of the polymorphism of the flagellar filament is discussed. 相似文献
17.
Bradyrhizobium japonicum is one of the soil bacteria that form nodules on soybean roots. The cell has two sets of flagellar systems, one thick flagellum and a few thin flagella, uniquely growing at subpolar positions. The thick flagellum appears to be semicoiled in morphology, and the thin flagella were in a tight-curly form as observed by dark-field microscopy. Flagellin genes were identified from the amino acid sequence of each flagellin. Flagellar genes for the thick flagellum are scattered into several clusters on the genome, while those genes for the thin flagellum are compactly organized in one cluster. Both types of flagella are powered by proton-driven motors. The swimming propulsion is supplied mainly by the thick flagellum. B. japonicum flagellar systems resemble the polar-lateral flagellar systems of Vibrio species but differ in several aspects. 相似文献
18.
Identification and sequence analysis of two related flagellin genes in Rhizobium meliloti. 总被引:28,自引:22,他引:6
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The genomic region that codes for the flagellin subunits of the complex flagellar filaments of Rhizobium meliloti was cloned and sequenced. Two structural genes, flaA and flaB, that encode 395- and 396-amino-acid polypeptides, respectively, were identified. These exhibit 87% sequence identity. The amino acid sequences of tryptic peptides suggest that both of these subunit proteins are represented in the flagellar filaments. The N-terminal methionine was absent from the mature flagellin subunits. Their derived primary structures show almost no relationship to flagellins from Escherichia coli, Salmonella typhimurium, or Bacillus subtilis but exhibit up to 60% similarity to the N- and C-terminal portions of flagellin from Caulobacter crescentus. It is suggested that the complex flagellar filaments of R. meliloti are unique in being assembled from heterodimers of two related flagellin subunits. The tandemly arranged flagellin genes were shown to be transcribed separately from unusual promoter sequences. 相似文献
19.
Sequence and characterization of Bacillus subtilis CheB, a homolog of Escherichia coli CheY, and its role in a different mechanism of chemotaxis 总被引:10,自引:0,他引:10
The Bacillus subtilis gene encoding CheB, which is homologous to Escherichia coli CheY, the regulator of flagellar rotation, has been cloned and sequenced. It has been verified, using a phage T7 expression system, by showing that a small protein, the same size as E. coli CheY, is actually made from this DNA. Despite the fact that the two proteins are 36% identical, with many highly conserved residues, they appear to play different roles. Unlike CheY null mutants, which swim smoothly, CheB null mutants tumble incessantly. However, a CheB point mutant swims smoothly, even in the presence of a plasmid bearing cheB, which restores the null mutants to wild type. Expression of CheB in wild type B. subtilis makes the cells exhibit more tumbling. Since both absence of CheB and presence of high levels of CheB cause tumbling, CheB appears to be required, in certain circumstances, for both smooth swimming and tumbling. Expression in wild type E. coli makes the cells smooth swimmers and strongly inhibits chemotaxis. 相似文献
20.
Termini of Salmonella flagellin are disordered and become organized upon polymerization into flagellar filament 总被引:5,自引:0,他引:5
The terminal regions of Salmonella flagellin are essential for polymerization to form the flagellar filament. It has recently been suggested, on the basis of results from circular dichroism spectroscopy and scanning calorimetry, that these regions are disordered in solution. We report here direct evidence for disorder and mobility in the terminal regions of flagellin using 400 MHz proton nuclear magnetic resonance (n.m.r.) spectroscopy. Comparison of the n.m.r. spectra of monomeric and polymeric flagellin shows that the terminal regions become organized when polymerized to form the filament. 相似文献