Work probability distribution in single molecule experiments

We derive and solve a differential equation satisfied by the probability distribution of the work done on a single biomolecule in a mechanical unzipping experiment. The unzipping is described as a thermally activated escape process in an energy landscape. The Jarzynski equality is recovered as an identity, independent of the pulling protocol. This approach allows one to evaluate easily, by numerical integration, the work distribution, once a few parameters of the energy landscape are known.Comment: To appear on EP

An Ising-Like model for protein mechanical unfolding

The mechanical unfolding of proteins is investigated by extending the Wako-Saito-Munoz-Eaton model, a simplified protein model with binary degrees of freedom, which has proved successful in describing the kinetics of protein folding. Such a model is generalized by including the effect of an external force, and its thermodynamics turns out to be exactly solvable. We consider two molecules, the 27th immunoglobulin domain of titin and protein PIN1. In the case of titin we determine equilibrium force-extension curves and study nonequilibrium phenomena in the frameworks of dynamic loading and force clamp protocols, verifying theoretical laws and finding the position of the kinetic barrier which hinders the unfolding of the molecule. The PIN1 molecule is used to check the possibility of computing the free energy landscape as a function of the molecule length by means of an extended form of the Jarzynski equality.Comment: 4 pages + appendi

Internal Motility in Stiffening Actin-Myosin Networks

We present a study on filamentous actin solutions containing heavy meromyosin subfragments of myosin II motor molecules. We focus on the viscoelastic phase behavior and internal dynamics of such networks during ATP depletion. Upon simultaneously using micro-rheology and fluorescence microscopy as complementary experimental tools, we find a sol-gel transition accompanied by a sudden onset of directed filament motion. We interpret the sol-gel transition in terms of myosin II enzymology, and suggest a "zipping" mechanism to explain the filament motion in the vicinity of the sol-gel transition.Comment: 4 pages, 3 figure

Rupture of multiple parallel molecular bonds under dynamic loading

Biological adhesion often involves several pairs of specific receptor-ligand molecules. Using rate equations, we study theoretically the rupture of such multiple parallel bonds under dynamic loading assisted by thermal activation. For a simple generic type of cooperativity, both the rupture time and force exhibit several different scaling regimes. The dependence of the rupture force on the number of bonds is predicted to be either linear, like a square root or logarithmic.Comment: 8 pages, 2 figure

Campylobacter jejuni motility integrates specialized cell shape, flagellar filament, and motor, to coordinate action of its opposed flagella

Campylobacter jejuni rotates a flagellum at each pole to swim through the viscous mucosa of its hosts’ gastrointestinal tracts. Despite their importance for host colonization, however, how C. jejuni coordinates rotation of these two opposing flagella is unclear. As well as their polar placement, C. jejuni’s flagella deviate from the norm of Enterobacteriaceae in other ways: their flagellar motors produce much higher torque and their flagellar filament is made of two different zones of two different flagellins. To understand how C. jejuni’s opposed motors coordinate, and what contribution these factors play in C. jejuni motility, we developed strains with flagella that could be fluorescently labeled, and observed them by high-speed video microscopy. We found that C. jejuni coordinates its dual flagella by wrapping the leading filament around the cell body during swimming in high-viscosity media and that its differentiated flagellar filament and helical body have evolved to facilitate this wrapped-mode swimming

Effect of Silicon Content on Carbide Precipitation and Low-Temperature Toughness of Pressure Vessel Steels

Cr – Mn – Mo – Ni pressure vessel steels containing 0.54 and 1.55% Si are studied. Metallographic and fractographic analyses of the steels after tempering at 650 and 700°C are performed. The impact toughness at – 30°C and the hardness of the steels are determined. The mass fraction of the carbide phase in the steels is computed with the help of the J-MatPro 4.0 software

Possible origins of macroscopic left-right asymmetry in organisms

I consider the microscopic mechanisms by which a particular left-right (L/R) asymmetry is generated at the organism level from the microscopic handedness of cytoskeletal molecules. In light of a fundamental symmetry principle, the typical pattern-formation mechanisms of diffusion plus regulation cannot implement the "right-hand rule"; at the microscopic level, the cell's cytoskeleton of chiral filaments seems always to be involved, usually in collective states driven by polymerization forces or molecular motors. It seems particularly easy for handedness to emerge in a shear or rotation in the background of an effectively two-dimensional system, such as the cell membrane or a layer of cells, as this requires no pre-existing axis apart from the layer normal. I detail a scenario involving actin/myosin layers in snails and in C. elegans, and also one about the microtubule layer in plant cells. I also survey the other examples that I am aware of, such as the emergence of handedness such as the emergence of handedness in neurons, in eukaryote cell motility, and in non-flagellated bacteria.Comment: 42 pages, 6 figures, resubmitted to J. Stat. Phys. special issue. Major rewrite, rearranged sections/subsections, new Fig 3 + 6, new physics in Sec 2.4 and 3.4.1, added Sec 5 and subsections of Sec

Ligand-Receptor Interactions

The formation and dissociation of specific noncovalent interactions between a variety of macromolecules play a crucial role in the function of biological systems. During the last few years, three main lines of research led to a dramatic improvement of our understanding of these important phenomena. First, combination of genetic engineering and X ray cristallography made available a simultaneous knowledg of the precise structure and affinity of series or related ligand-receptor systems differing by a few well-defined atoms. Second, improvement of computer power and simulation techniques allowed extended exploration of the interaction of realistic macromolecules. Third, simultaneous development of a variety of techniques based on atomic force microscopy, hydrodynamic flow, biomembrane probes, optical tweezers, magnetic fields or flexible transducers yielded direct experimental information of the behavior of single ligand receptor bonds. At the same time, investigation of well defined cellular models raised the interest of biologists to the kinetic and mechanical properties of cell membrane receptors. The aim of this review is to give a description of these advances that benefitted from a largely multidisciplinar approach

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 “strokes”; the “power stroke” is the force-generating swinging of the myosin light chain–binding “neck” domain relative to the motor domain “head” while bound to actin; the “recovery stroke” is the necessary initial motion that primes, or “cocks,” myosin while detached from actin. Myosin Va is a processive dimer that steps unidirectionally along actin following a “hand over hand” mechanism in which the trailing head detaches and steps forward ∼72 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 ∼40 s, until ATP hydrolysis products are released. Angle distributions indicate that the post-recovery stroke conformation is stabilized by ≥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

Detection of peptide-specific CTL-precursors in peripheral blood lymphocytes of cancer patients

Development of therapeutic vaccines is one of the major areas of tumour immunotherapy today. However, clinical trials of peptide-based cancer vaccines have rarely resulted in tumour regression. This failure might be due to an insufficient induction of cytotoxic T lymphocytes in the current regimes, in which cytotoxic T lymphocytes-precursors in pre-vaccination peripheral blood mononuclear cells are not measured. Initiation of immune-boosting through vaccination could be better than that of immune-priming with regard to induction of prompt and strong immunity. If this is also the case for therapeutic vaccines, pre-vaccination measurement of peptide-specific cytotoxic T lymphocytes-precursors will be important. In the present study, we investigated whether cytotoxic T lymphocytes-precursors reacting to 28 kinds of peptides of vaccine candidates (13 and 15 peptides for HLA-A24+ and HLA-A2+ patients, respectively) were detectable in pre-vaccination peripheral blood mononuclear cells of 80 cancer patients. Peptide-specific cytotoxic T lymphocytes-precursors were found to be detectable in peripheral blood mononuclear cells of the majority of cancer patients (57 out of 80 cases, 71%). The mean numbers of positive peptides were 2.0 peptides per positive case. Peripheral blood mononuclear cells incubated with positive peptides, not with negative peptides, showed significant levels of HLA-class-I-restricted cytotoxicity to cancer cells. The profiles of positive peptides entirely varied among patients, and were not influenced by the cancer origin. These results may provide a scientific basis for the development of a new approach to cancer immunotherapy, e.g.) cytotoxic T lymphocytes-precursor-oriented peptide vaccine