scholarly journals On Torque and Tumbling in Swimming Escherichia coli

2006 ◽  
Vol 189 (5) ◽  
pp. 1756-1764 ◽  
Author(s):  
Nicholas C. Darnton ◽  
Linda Turner ◽  
Svetlana Rojevsky ◽  
Howard C. Berg
Keyword(s):  
Escherichia Coli ◽  
High Speed ◽  
Polymorphic Transformations ◽  
Charge Coupled Device ◽  
Motor Torque ◽  
Rotation Rates ◽  
Applied Torque ◽  
Resistive Force Theory ◽  
Rotary Motor ◽  
Single Flagellum

ABSTRACT Bacteria swim by rotating long thin helical filaments, each driven at its base by a reversible rotary motor. When the motors of peritrichous cells turn counterclockwise (CCW), their filaments form bundles that drive the cells forward. We imaged fluorescently labeled cells of Escherichia coli with a high-speed charge-coupled-device camera (500 frames/s) and measured swimming speeds, rotation rates of cell bodies, and rotation rates of flagellar bundles. Using cells stuck to glass, we studied individual filaments, stopping their rotation by exposing the cells to high-intensity light. From these measurements we calculated approximate values for bundle torque and thrust and body torque and drag, and we estimated the filament stiffness. For both immobilized and swimming cells, the motor torque, as estimated using resistive force theory, was significantly lower than the motor torque reported previously. Also, a bundle of several flagella produced little more torque than a single flagellum produced. Motors driving individual filaments frequently changed directions of rotation. Usually, but not always, this led to a change in the handedness of the filament, which went through a sequence of polymorphic transformations, from normal to semicoiled to curly 1 and then, when the motor again spun CCW, back to normal. Motor reversals were necessary, although not always sufficient, to cause changes in filament chirality. Polymorphic transformations among helices having the same handedness occurred without changes in the sign of the applied torque.

scholarly journals Site-Directed Cross-Linking Identifies the Stator-Rotor Interaction Surfaces in a Hybrid Bacterial Flagellar Motor

2021 ◽  
Vol 203 (9) ◽  
Author(s):  
Hiroyuki Terashima ◽  
Seiji Kojima ◽  
Michio Homma
Keyword(s):  
Escherichia Coli ◽  
Cross Linking ◽  
Physical Interaction ◽  
Flagellar Motor ◽  
Intact Cells ◽  
Content Type ◽  
Bacterial Flagellum ◽  
Cross Links ◽  
Bacterial Flagellar Motor ◽  
Rotary Motor

ABSTRACT The bacterial flagellum is the motility organelle powered by a rotary motor. The rotor and stator elements of the motor are located in the cytoplasmic membrane and cytoplasm. The stator units assemble around the rotor, and an ion flux (typically H+ or Na+) conducted through a channel of the stator induces conformational changes that generate rotor torque. Electrostatic interactions between the stator protein PomA in Vibrio (MotA in Escherichia coli) and the rotor protein FliG have been shown by genetic analyses but have not been demonstrated biochemically. Here, we used site-directed photo-cross-linking and disulfide cross-linking to provide direct evidence for the interaction. We introduced a UV-reactive amino acid, p-benzoyl-l-phenylalanine (pBPA), into the cytoplasmic region of PomA or the C-terminal region of FliG in intact cells. After UV irradiation, pBPA inserted at a number of positions in PomA and formed a cross-link with FliG. PomA residue K89 gave the highest yield of cross-links, suggesting that it is the PomA residue nearest to FliG. UV-induced cross-linking stopped motor rotation, and the isolated hook-basal body contained the cross-linked products. pBPA inserted to replace residue R281 or D288 in FliG formed cross-links with the Escherichia coli stator protein, MotA. A cysteine residue introduced in place of PomA K89 formed disulfide cross-links with cysteine inserted in place of FliG residues R281 and D288 and some other flanking positions. These results provide the first demonstration of direct physical interaction between specific residues in FliG and PomA/MotA. IMPORTANCE The bacterial flagellum is a unique organelle that functions as a rotary motor. The interaction between the stator and rotor is indispensable for stator assembly into the motor and the generation of motor torque. However, the interface of the stator-rotor interaction has only been defined by mutational analysis. Here, we detected the stator-rotor interaction using site-directed photo-cross-linking and disulfide cross-linking approaches. We identified several residues in the PomA stator, especially K89, that are in close proximity to the rotor. Moreover, we identified several pairs of stator and rotor residues that interact. This study directly demonstrates the nature of the stator-rotor interaction and suggests how stator units assemble around the rotor and generate torque in the bacterial flagellar motor.

scholarly journals Dynamic mechanisms of CRISPR interference by Escherichia coli CRISPR-Cas3

2021 ◽  
Author(s):  
Kazuto Yoshimi ◽  
Kohei TAKESHITA ◽  
Noriyuki Kodera ◽  
Satomi Shibumura ◽  
Yuko Yamauchi ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
High Speed ◽  
Dna Helicase ◽  
Type I ◽  
Loop Formation ◽  
Force Microscopy ◽  
Atomic Force ◽  
Target Strand ◽  
Target Dna

Type I CRISPR-Cas3 uses an RNA-guided multi Cas-protein complex, Cascade, which detects and degrades foreign nucleic acids via the helicase-nuclease Cas3 protein. Despite many studies using cryoEM and smFRET, the precise mechanism of Cas3-mediated cleavage and degradation of target DNA remains elusive. Here we reconstitute the CRISPR-Cas3 system in vitro to show how the Escherichia coli Cas3 (EcoCas3) with EcoCascade exhibits collateral non-specific ssDNA cleavage and target specific DNA degradation. Partial binding of EcoCascade to target DNA with tolerated mismatches within the spacer sequence, but not the PAM, elicits collateral ssDNA cleavage activity of recruited EcoCas3. Conversely, stable binding with complete R-loop formation drives EcoCas3 to nick the non-target strand (NTS) in the bound DNA. Helicase-dependent unwinding then combines with trans ssDNA cleavage of the target strand and repetitive cis cleavage of the NTS to degrade the target dsDNA substrate. High-speed atomic force microscopy demonstrates that EcoCas3 bound to EcoCascade repeatedly reels and releases the target DNA, followed by target fragmentation. Together, these results provide a revised model for collateral ssDNA cleavage and target dsDNA degradation by CRISPR-Cas3, furthering understanding of type I CRISPR priming and interference and informing future genome editing tools.

Sensorimotor aspects of high-speed artificial gravity: III. Sensorimotor adaptation

2003 ◽  
Vol 12 (5-6) ◽  
pp. 291-299
Author(s):  
Paul DiZio ◽  
James R. Lackner
Keyword(s):  
Side Effects ◽  
Coriolis Force ◽  
High Speed ◽  
Motor Adaptation ◽  
Vestibular Stimulation ◽  
Head Movements ◽  
Artificial Gravity ◽  
Slow Rotation ◽  
Movement Trajectory ◽  
Rotation Rates

As a countermeasure to the debilitating physiological effects of weightlessness, astronauts could live continuously in an artificial gravity environment created by slow rotation of an entire spacecraft or be exposed to brief daily "doses" in a short radius centrifuge housed within a non-rotating spacecraft. A potential drawback to both approaches is that head movements made during rotation may be disorienting and nauseogenic. These side effects are more severe at higher rotation rates, especially upon first exposure. Head movements during rotation generate aberrant vestibular stimulation and Coriolis force perturbations of the head-neck motor system. This article reviews our progress toward distinguishing vestibular and motor factors in side effects of rotation, and presents new data concerning the rates of rotation up to which adaptation is possible. We have studied subjects pointing to targets during constant velocity rotation, because these movements generate Coriolis motor perturbations of the arm but do not involve unusual vestibular stimulation. Initially, reaching paths and endpoints are deviated in the direction of the transient lateral Coriolis forces generated. With practice, subjects soon move in straighter paths and land on target once more. If sight of the arm is permitted, adaptation is more rapid than in darkness. Initial arm movement trajectory and endpoint deviations are proportional to Coriolis force magnitude over a range of rotation speeds from 5 to 20 rpm, and there is rapid, complete motor adaptation at all speeds. These new results indicate that motor adaptation to high rotation rates is possible. Coriolis force perturbations of head movements also occur in a rotating environment but adaptation gradually develops over the course of many head movements.

Hygienic Effects of Trimming and Washing Operations in a Beef-Carcass-Dressing Process†

1996 ◽  
Vol 59 (6) ◽  
pp. 666-669 ◽  
Author(s):  
C. O. GILL ◽  
M. BADONI ◽  
T. JONES
Keyword(s):  
Escherichia Coli ◽  
Standard Deviation ◽  
High Speed ◽  
Effective Means ◽  
Arithmetic Mean ◽  
Single Sample ◽  
Aerobic Bacteria ◽  
E Coli ◽  
Beef Carcasses ◽  
The Mean

Swab samples were obtained from the surfaces of randomly selected beef carcasses passing through a high-speed dressing process. A single sample was obtained from a randomly selected site on the surface of each selected carcass. Fifty such samples were collected at each of four stages in the process. The aerobic bacteria, coliforms, and Escherichia coli recovered from each sample were enumerated. Values for the mean log units and standard deviations of each set of 50 log values were calculated on the assumption that the log values were normally distributed. The log of the arithmetic mean was estimated from the mean log and standard deviation values for each set. The results show that the average numbers of E. coli, coliforms, and aerobic bacteria which are deposited on carcasses during skinning and evisceration are not reduced by trimming, and that washing approximately halves the average numbers of those bacteria on carcasses. It is concluded that commercial trimming and washing operations are not effective means of decontaminating beef carcasses.

Real-time lock-in imaging by a newly developed high-speed image-processing charge coupled device video camera

2003 ◽  
Vol 74 (3) ◽  
pp. 1393-1396 ◽  
Author(s):  
Kentarou Nishikata ◽  
Yoshihide Kimura ◽  
Yoshizo Takai ◽  
Takashi Ikuta ◽  
Ryuichi Shimizu
Keyword(s):  
Image Processing ◽  
Real Time ◽  
High Speed ◽  
Video Camera ◽  
Charge Coupled Device ◽  
Lock In ◽  
High Speed Image Processing

scholarly journals Dynamics of flagellar force generated by a hyperactivated spermatozoon

Reproduction ◽  
2011 ◽  
Vol 142 (3) ◽  
pp. 409-415 ◽  
Author(s):  
Sumio Ishijima
Keyword(s):  
Zona Pellucida ◽  
High Speed ◽  
Beat Frequency ◽  
Transverse Force ◽  
Sperm Head ◽  
Propulsive Force ◽  
Resistive Force ◽  
High Speed Video ◽  
Sperm Penetration ◽  
Resistive Force Theory

The flagellar force generated by a hyperactivated monkey spermatozoon was evaluated using the resistive force theory applied to the activated (nonhyperactivated) and hyperactivated flagellar waves that were obtained using high-speed video microscopy and digital image processing in order to clarify the mechanism of sperm penetration through the zona pellucida. No difference in the maximum propulsive force, which was parallel to the longitudinal sperm head axis, was found between the activated and hyperactivated spermatozoa. The maximum transverse force (45 pN), which was perpendicular to the longitudinal sperm head axis, of the hyperactivated spermatozoon was ∼2.5 times its propulsive force. As the beat frequency of the flagellar beating remarkably decreased during the hyperactivation, the slowly oscillating transverse force (5 Hz) by the hyperactivated spermatozoon seems to be most effective for sperm penetration through the zona pellucida.

High Speed Document Scanner Using Charge Coupled Device (CCD) Image Sensors

1983 ◽  
Author(s):  
Masahiro Mori ◽  
Isao Kondo ◽  
Masakatsu Horie
Keyword(s):  
High Speed ◽  
Image Sensors ◽  
Charge Coupled Device ◽  
Ccd Image
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