369 research outputs found
Spatiotemporal relationships among early events of fertilization in sea urchin eggs revealed by multiview microscopy
Four early events of egg fertilization, changes in intracellular calcium concentration and intracellular pH, reorientation of the surface membrane, and the elevation of the fertilization envelope, were imaged in real time and in pairs in single sea urchin eggs. The paired imaging allowed the correlation of the four events spatially and temporally. Three of them propagated as waves starting at the sperm entry site. The earliest was the calcium wave, visualized with fluorescent indicator dyes. After a delay of 10 s there followed a large decrease in the fluorescence polarization of membrane-bound dyes, which we interpret as arising from membrane reorientation as a result of cortical granule exocytosis and microvillar elongation. With a further delay of 15 s the fertilization envelope was seen to rise in transmitted light. All three waves propagated with similar velocities of approximately 10 microns/s, supporting the view that calcium triggers the latter two events. The fluorescence polarization changed in two steps with a clear pause of 10â20 s in between. The second step, which also propagated as wave, reflects either further elongation of microvilli or straightening of irregular microvilli. This second step was abolished by cytochalasin B and was coincident with an increase in cytoplasmic pH, suggesting that pH-induced actin reorganization may play a role. The cytoplasmic alkalinization, imaged with a fluorescent probe, was quite different from the other events in that it took place homogeneously throughout the egg and slowly (over 100 s). Apparently, the alkalinization is not on a direct downstream pathway of calcium origin. An opposing possibility, that the alkalinization may in fact be triggered by the traveling calcium wave, is also discussed
Molecular Chemical Engines: Pseudo-Static Processes and the Mechanism of Energy Transduction
We propose a simple theoretical model for a molecular chemical engine that
catalyzes a chemical reaction and converts the free energy released by the
reaction into mechanical work. Binding and unbinding processes of reactant and
product molecules to and from the engine are explicitly taken into account. The
work delivered by the engine is calculated analytically for infinitely slow
(``pseudo-static'') processes, which can be reversible (quasi-static) or
irreversible, controlled by an external agent. It is shown that the work larger
than the maximum value limited by the second law of thermodynamics can be
obtained in a single cycle of operation by chance, although the statistical
average of the work never exceeds this limit and the maximum work is delivered
if the process is reversible. The mechanism of the energy transductionis also
discussed.Comment: 8 pages, 3 figues, submitted to J. Phys. Soc. Jp
Efficiency of Energy Transduction in a Molecular Chemical Engine
A simple model of the two-state ratchet type is proposed for molecular
chemical engines that convert chemical free energy into mechanical work and
vice versa. The engine works by catalyzing a chemical reaction and turning a
rotor. Analytical expressions are obtained for the dependences of rotation and
reaction rates on the concentrations of reactant and product molecules, from
which the performance of the engine is analyzed. In particular, the efficiency
of energy transduction is discussed in some detail.Comment: 4 pages, 4 fugures; title modified, figures 2 and 3 modified, content
changed (pages 1 and 4, mainly), references adde
The ATP-waiting conformation of rotating F1-ATPase revealed by single-pair fluorescence resonance energy transfer
F1-ATPase is an ATP-driven rotary motor in which a rod-shaped gamma subunit rotates inside a cylinder made of alpha3beta3 subunits. To elucidate the conformations of rotating F1, we measured fluorescence resonance energy transfer (FRET) between a donor on one of the three betas and an acceptor on gamma in single F1 molecules. The yield of FRET changed stepwise at low ATP concentrations, reflecting the stepwise rotation of gamma. In the ATP-waiting state, the FRET yields indicated a gamma position approximately 40 degrees counterclockwise (= direction of rotation) from that in the crystal structures of mitochondrial F1, suggesting that the crystal structures mimic a metastable state before product release
Extrapolation-CAM Theory for Critical Exponents
By intentionally underestimating the rate of convergence of
exact-diagonalization values for the mass or energy gaps of finite systems, we
form families of sequences of gap estimates. The gap estimates cross zero with
generically nonzero linear terms in their Taylor expansions, so that
for each member of these sequences of estimates. Thus, the Coherent Anomaly
Method can be used to determine . Our freedom in deciding exactly how to
underestimate the convergence allows us to choose the sequence that displays
the clearest coherent anomaly. We demonstrate this approach on the
two-dimensional ferromagnetic Ising model, for which . We also use it
on the three-dimensional ferromagnetic Ising model, finding , in good agreement with other estimates.Comment: 21 pages, Submitted to Journal of Physics A; new section added
discussing rate of convergence and relation to Finite-Size Scalin
Direct Observation of the Myosin Va Recovery Stroke That Contributes to Unidirectional Stepping along Actin
Myosins are ATP-driven linear molecular motors that work as cellular force
generators, transporters, and force sensors. These functions are driven by
large-scale nucleotide-dependent conformational changes, termed
âstrokesâ; the âpower strokeâ is the force-generating
swinging of the myosin light chainâbinding âneckâ domain
relative to the motor domain âheadâ while bound to actin; the
ârecovery strokeâ is the necessary initial motion that primes, or
âcocks,â myosin while detached from actin. Myosin Va is a processive
dimer that steps unidirectionally along actin following a âhand over
handâ mechanism in which the trailing head detaches and steps forward
âŒ72 nm. Despite large rotational Brownian motion of the detached head about
a free joint adjoining the two necks, unidirectional stepping is achieved, in
part by the power stroke of the attached head that moves the joint forward.
However, the power stroke alone cannot fully account for preferential forward
site binding since the orientation and angle stability of the detached head,
which is determined by the properties of the recovery stroke, dictate actin
binding site accessibility. Here, we directly observe the recovery stroke
dynamics and fluctuations of myosin Va using a novel, transient caged
ATP-controlling system that maintains constant ATP levels through stepwise
UV-pulse sequences of varying intensity. We immobilized the neck of monomeric
myosin Va on a surface and observed real time motions of bead(s) attached
site-specifically to the head. ATP induces a transient swing of the neck to the
post-recovery stroke conformation, where it remains for âŒ40 s, until ATP
hydrolysis products are released. Angle distributions indicate that the
post-recovery stroke conformation is stabilized by â„5
kBT of energy. The high kinetic
and energetic stability of the post-recovery stroke conformation favors
preferential binding of the detached head to a forward site 72 nm away. Thus,
the recovery stroke contributes to unidirectional stepping of myosin Va
Exploring metabolic responses of potato tissue induced by electric pulses
In this study, we investigated the metabolic
responses of potato tissue induced by pulsed electric field
(PEF). Potato tissue was subjected to field strengths ranging
from 30 to 500 V/cm, with a single rectangular pulse of 10 ÎŒs,
100 ÎŒs, or 1 ms. Metabolic responses were monitored using
isothermal calorimetry, changes on electrical resistance during
the delivery of the pulse, as well as impedance measurements.
Our results show that the metabolic response involves oxygen
consuming pathways as well as other unidentified events that
are shown to be insensitive to metabolic inhibitors such as
KCN and sodium azide. The metabolic response is strongly
dependent on pulsing conditions and is independent of the
total permeabilization achieved by the pulse. Evidence shows
that calorimetry is a simple and powerful method for
exploring conditions for metabolic stimulation, providing
information on metabolic responses that can not be obtained
from electrical measurements. This study set the basis for
further investigations on defense-related consequences of
PEF-induced stress.Sparbanksstiftelsen FĂ€rs & Frosta (Sweden).Fundação para a CiĂȘncia e a Tecnologia (FCT).Lund University (Sweden).Department of Cell and Organism Biology; Department of Plant Biochemistry
Electric Field Exposure Triggers and Guides Formation of Pseudopod-Like Blebs in U937 Monocytes
We describe a new phenomenon of anodotropic pseudopod-like blebbing in U937 cells stimulated by nanosecond pulsed electric field (nsPEF). In contrast to regular, round-shaped blebs, which are often seen in response to cell damage, pseudopod-like blebs (PLBs) formed as longitudinal membrane protrusions toward anode. PLB length could exceed the cell diameter in 2 min of exposure to 60-ns, 10-kV/cm pulses delivered at 10-20 Hz. Both PLBs and round-shaped nsPEF-induced blebs could be efficiently inhibited by partial isosmotic replacement of bath NaCl for a larger solute (sucrose), thereby pointing to the colloid-osmotic water uptake as the principal driving force for bleb formation. In contrast to round-shaped blebs, PLBs retracted within several minutes after exposure. Cells treated with 1 nM of the actin polymerization blocker cytochalasin D were unable to form PLBs and instead produced stationary, spherical blebs with no elongation or retraction capacity. Live cell fluorescent actin tagging showed that during elongation actin promptly entered the PLB interior, forming bleb cortex and scaffold, which was not seen in stationary blebs. Overall, PLB formation was governed by both passive (physicochemical) effects of membrane permeabilization and active cytoskeleton assembly in the living cell. To a certain extent, PLB mimics the membrane extension in the process of cell migration and can be employed as a nonchemical model for studies of cytomechanics, membrane-cytoskeleton interaction and cell motility
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