Helical Flagellar Filaments Research Articles

Resurrection of the flagellar rotary motor near zero load

Flagellated bacteria, such as Escherichia coli, are propelled by helical flagellar filaments, each driven at its base by a reversible rotary motor, powered by a transmembrane proton flux. Torque is generated by the interaction of stator proteins, MotA and MotB, with a rotor protein FliG. The physiology of the motor has been studied extensively in the regime of relatively high load and low speed, where it appears to operate close to thermodynamic equilibrium. Here, we describe an assay that allows systematic study of the motor near zero load, where proton translocation and movement of mechanical components are rate limiting. Sixty-nanometer-diameter gold spheres were attached to hooks of cells lacking flagellar filaments, and light scattered from a sphere was monitored at the image plane of a microscope through a small pinhole. Paralyzed motors of cells carrying a motA point mutation were resurrected at 23 degrees C by expression of wild-type MotA, and speeds jumped from zero to a maximum value ( approximately 300 Hz) in one step. Thus, near zero load, the speed of the motor is independent of the number of torque-generating units. Evidently, the units act independently (they do not interfere with one another), and there are no intervals during which a second unit can add to the speed generated by the first (the duty ratio is close to 1).

Model for polymorphic transitions in bacterial flagella

Many bacteria use rotating helical flagellar filaments to swim. The filaments undergo polymorphic transformations in which the helical pitch and radius change abruptly. These transformations arise in response to mechanical loading, changes in solution temperature and ionic strength, and point substitutions in the amino acid sequence of the protein subunits that make up the filament. To explain polymorphism, we propose a coarse-grained continuum rod theory based on the quaternary structure of the filament. The model has two molecular switches. The first is a double-well potential for the extension of a protofilament, which is one of the 11 almost longitudinal columns of subunits. Curved filament shapes occur in the model when there is a mismatch strain, i.e., when intersubunit bonds in the inner core of the filament prefer a subunit spacing which is intermediate between the two spacings favored by the double-well potential. The second switch is a double-well potential for twist, due to lateral interactions between neighboring protofilaments. Cooperative interactions between neighboring subunits within a protofilament are necessary to ensure the uniqueness of helical ground states. We calculate a phase diagram for filament shapes and the response of a filament to external moment and force.

Role of body rotation in bacterial flagellar bundling.

In bacterial chemotaxis, E. coli cells drift up chemical gradients by a series of runs and tumbles. Runs are periods of directed swimming, and tumbles are abrupt changes in swimming direction. Near the beginning of each run, the rotating helical flagellar filaments that propel the cell form a bundle. Using resistive-force theory, we show that the counterrotation of the cell body necessary for torque balance is sufficient to wrap the filaments into a bundle, even in the absence of the swirling flows produced by each individual filament.

Envelope structure of Synechococcus sp. WH8113, a nonflagellated swimming cyanobacterium

BackgroundMany bacteria swim by rotating helical flagellar filaments [1]. Waterbury et al. [15] discovered an exception, strains of the cyanobacterium Synechococcus that swim without flagella or visible changes in shape. Other species of cyanobacteria glide on surfaces [2,7]. The hypothesis that Synechococcus might swim using traveling surface waves [6,13] prompted this investigation.ResultsUsing quick-freeze electron microscopy, we have identified a crystalline surface layer that encloses the outer membrane of the motile strain Synechococcus sp. WH8113, the components of which are arranged in a rhomboid lattice. Spicules emerge in profusion from the layer and extend up to 150 nm into the surrounding fluid. These spicules also send extensions inwards to the inner cell membrane where motility is powered by an ion-motive force [17].ConclusionThe envelope structure of Synechococcus sp. WH8113 provides new constraints on its motile mechanism. The spicules are well positioned to transduce energy at the cell membrane into mechanical work at the cell surface. One model is that an unidentified motor embedded in the cell membrane utilizes the spicules as oars to generate a traveling wave external to the surface layer in the manner of ciliated eukaryotes.

102高速回転している細菌べん毛のらせんピッチの観測

Many kind of bacteria swim in aqueous solution by rotating their helical flagellar filaments. Each filament is driven by a rotary motor (flagellar motor) embedded in the cytoplasmic membrane and can rotate very fast. The rotation speed reaches from 100 to more than 1000 r/sec. The possible deformation of such fast rotating flagellar filaments has not yet been observed because of difficulties mainly caused by their extremely small size. We tried to observe the deformation of the flagellar helices of swimming Vibrio alginolyticus cells with laser dark-field microscope. The laser dark-field image was recorded by using an image intensified CCD camera equipped with the electrical shutter. The change in the helical pitch was observed to be rather small, around 10%.

Cloning and characterization of motY, a gene coding for a component of the sodium-driven flagellar motor in Vibrio alginolyticus.

The bacterial flagellar motor is a molecular machine that couples proton or sodium influx to force generation for driving rotation of the helical flagellar filament. In this study, we cloned a gene (motY) encoding a component of the sodium-driven polar flagellar motor in Vibrio alginolyticus. Nucleotide sequence analysis revealed that the gene encodes a 293-amino-acid polypeptide with a single putative transmembrane segment that is very similar (94.5% identity) to the recently described MotY of V. parahaemolyticus. Their C-terminal domains were similar to the C-terminal domains of many peptidoglycan-interacting proteins, e.g., Escherichia coli MotB and OmpA, suggesting that MotY may interact with peptidoglycan for anchoring the motor. By using the lac promoter-repressor system, motY expression was controlled in V. alginolyticus cells. Swimming ability increased with increasing concentrations of the inducer isopropyl-beta-D-thiogalactopyranoside, and the swimming fraction increased after induction. These results are consistent with the notion that MotY is a component of the force-generating unit. V. alginolyticus motY complemented the motY mutation of V. parahaemolyticus. However, motY appeared to lack a region corresponding to the proposed motY promoter of V. parahaemolyticus. Instead, sequences similar to the sigma54 consensus were found in the upstream regions of both species. We propose that they are transcribed from the sigma54 -specific promoters.

Introduction to bacterial motility and chemotaxis

Bacteria swim by rotating semirigid, left-handed helical flagellar filaments; counterclockwise (CCW) rotation produces straight swims, known as "runs," and clockwise (CW) rotation generates abrupt changes in direction, known as "tumbles." As a cell moves through its environment, alternately running and tumbling, it detects spatial gradients of attractants and repellents by making temporal comparisons of their concentration. These chemicals bind to receptors in the cell envelope to modulate the activity of the chemotactic signal transducers, proteins that span the cytoplasmic membrane. Signals generated by the transducers control the motion of the flagella to promote migration up attractant gradients and down repellent gradients. Chemotactic adaptation, accomplished by methylation-demethylation of the transducers, cancels out these signals. Adaptation is an essential component of the "memory" that allows bacteria to use a temporal mechanism to detect spatial gradients. Both signaling and adaptation are mediated by changes in the level of phosphorylation of several cytoplasmic chemotaxis (Che) proteins. The activity of the transducers regulates the rate of autophosphorylation of the CheA protein, which then passes the phosphate on to other proteins. In particular, phosphorylated CheY protein controls the frequency of tumbling because it promotes CW flagellar rotation, and the CheB esterase modulates adaptation because its nonphosphorylated form removes methyl groups from the transducers much more slowly than its phosphorylated form.

Na+-driven bacterial flagellar motors.

Bacterial flagellar motors are the reversible rotary engine which propels the cell by rotating a helical flagellar filament as a screw propeller. The motors are embedded in the cytoplasmic membrane, and the energy for rotation is supplied by the electrochemical potential of specific ions across the membrane. Thus, the analysis of motor rotation at the molecular level is linked to an understanding of how the living system converts chemical energy into mechanical work. Based on the coupling ions, the motors are divided into two types; one is the H+-driven type found in neutrophiles such as Bacillus subtilis and Escherichia coli and the other is the Na+-driven type found in alkalophilic Bacillus and marine Vibrio. In this review, we summarize the current status of research on the rotation mechanism of the Na+-driven flagellar motors, which introduces several new aspects in the analysis.

The stall torque of the bacterial flagellar motor

The bacterial flagellar motor couples the flow of protons across the cytoplasmic membrane to the rotation of a helical flagellar filament. Using tethered cells, we have measured the stall torque required to block this rotation and compared it with the torque of the running motor over a wide range of values of proton-motive force and pH. The stall torque and the running torque vary identically: both appear to saturate at large values of the proton-motive force and both decrease at low or high pH. This suggests that up to speeds of approximately 5 Hz the operation of the motor is not limited by the mobility of its internal components or the rates of proton transfer reactions coupled to flagellar rotation.

A Phenomenological Theory of Polymorphism of Salmonella Flagella. I. A Model

A model theory has been proposed, which explains the polymorphism of Salmonella flagella observed by Asakura. Relation between the overall shapes of the helical flagellar filaments and the shapes of their constituents, i.e. , the flagellin monomer units is derived with the aid of two parameters, the curvature and the torsion of the helix. The model proposed is based upon the assumptions that the flagellin molecules in the flagellum assume two states R and T depending upon circumstances, as discussed by Asakura, that the flagellar filament consists of eight strands of the flagellin monomer units, that the five polymorphs have one, three, five, seven and eight T-type strands, respectively. Agreement is obtained between the observed and calculated values of curvatures and torsions of the helical flagellar filaments, and the parameters are determined, which characterize the shapes of the flagellin monomer units.