We report a simple method using semiconductor quantum dots (QDs) to track the motion of intracellular proteins with a high sensitivity. We characterized the in vivo motion of individual QD-tagged kinesin motors in living HeLa cells. Single-molecule measurements provided important parameters of the motor, such as its velocity and processivity, as well as an estimate of the force necessary to carry a QD. Our measurements demonstrate the importance of single-molecule experiments in the investigation of intracellular transport as well as the potential of single quantum-dot imaging for the study of important processes such as cellular trafficking, cell polarization, and division.

We report the tracking of single myosin V molecules in their natural environment, the cell. Myosin V molecules, labeled with quantum dots, are introduced into the cytoplasm of living HeLa cells and their motion is recorded at the single molecule level with high spatial and temporal resolution. We perform an intracellular measurement of key parameters of this molecular transporter: velocity, processivity, step size, and dwell time. Our experiments bridge the gap between in vitro single molecule assays and the indirect measurements of the motor features deduced from the tracking of organelles in live cells.

We demonstrate, for the first time, the large electromechanical response in nematic liquid crystalline elastomers filled with a very low (∼ 0.01%) concentration of carbon nanotubes, aligned along the nematic director at preparation. The nanotubes create a very large effective dielectric anisotropy of the composite. Their local field-induced torque is transmitted to the rubber-elastic network and is registered as the exerted uniaxial stress of order ∼ 1 kPa in response to a constant field of order ∼ 1 MV/m. We investigate the dependence of the effect on field strength, nanotube concentration and reproducibility under multiple field-on and -off cycles.The results indicate the potential of the nanotube-nematic elastomer composites as electrically driven actuators.

During gastrulation, dramatic movements rearrange cells into three germ layers expanded over the entire embryo [1-3]. In fish, both endoderm and mesoderm are specified as a belt at the embryo margin. Mesodermal layer expansion is achieved through the combination of two directed migrations. The outer ring of precursors moves toward the vegetal pole and continuously seeds mesodermal cells inside the embryo, which then reverse their movement in the direction of the animal pole [3-6]. Unlike mesoderm, endodermal cells internalize at once and must therefore adopt a different strategy to expand over the embryo [7, 8]. With live imaging of YFP-expressing zebrafish endodermal cells, we demonstrate that in contrast to mesoderm, internalized endodermal cells display a nonoriented/noncoordinated movement fit by a random walk that rapidly disperses them over the yolk surface. Transplantation experiments reveal that this behaviour is largely cell autonomous, induced by TGF-beta/Nodal, and dependent on the downstream effector Casanova. At midgastrulation, endodermal cells switch to a convergence movement. We demonstrate that this switch is triggered by environmental cues. These results uncover random walk as a novel Nodal-induced gastrulation movement and as an efficient strategy to transform a localized cell group into a layer expanded over the embryo.

Molecular chirality, and the chiral symmetry breaking of resulting macroscopic phases, can be topologically imprinted and manipulated by cross-linking and swelling of polymer networks. We present a new experimental approach to stereo-specific separation of chiral isomers by using a cholesteric elastomer in which a helical director distribution has been topologically imprinted by cross-linking. This makes the material unusual in that is has a strong phase chirality, but no molecular chirality at all; we study the nature and parameters controlling the twist-untwist transition. Adding a racemic mixture to the imprinted network results in selective swelling by only the component of "correct" handedness. We investigate the capacity of demixing in a racemic environment, which depends on network parameters and the underlying nematic order.

By combining dynamic mechanical and optical measurements in probing the internal structure of a biopolymer network (gelatin gel), we studied the quasi-equilibrium evolution of helical content as a function of the applied stress. Assuming that the net optical activity is proportional to the concentration of secondary helices of collagen chains, and assuming that affine mechanical deformation, we find a nonmonotonic relationship between the helical domains and an imposed deformation. The results are in qualitative agreement with theoretical predictions of ␣-helices induced by chain end-to-end stretching, and give a consistent picture of mechanically stimulated helix-coil transition in networks of denatured polypeptides.collagen ͉ helix ͉ optical rotation ͉ gelatin ͉ chirality I n recent years, a new field has emerged at the interface of physics and biology, aiming to explore structure and responses at molecular-length scales. Many single-molecule experiments have been performed to measure forces generated by biopolymers and their response to applied extension forces. The now classical work on DNA stretching (1) is just one of a number of significant recent advances in this field. By monitoring the response of a single molecule to pulling and twisting its ends [using atomic force microscopy (AFM), and magnetic trap and optical tweezer methods (2, 3)], one can probe the question of how chiral biopolymers are held in their native state, and their pathways of folding and unfolding. However, although the current techniques used in single-molecule force experiments reveal spectacular force-extension curves, they give little direct information about the structural transitions that occur on extension of these chains. Theoretical modeling of the behavior of single biopolymer molecules on extension has been hampered by this lack of information concerning structural changes. Using a new macroscopic approach of combining mechanical and optical methods in probing the internal structure of the biopolymer network, we found a direct relationship between helical domains and an externally imposed end-to-end distance of chains.Certain homopolypeptides form regular ␣-helices under appropriate conditions. In this case, the molecular configurations are well understood and are described according to the ZimmBragg model (4) [and its many modifications (5)], which assumes that each chain segment has access to only two conformational states, the random-coil state and the helical state. The average helical fraction of the chain can then be calculated for any number of model interactions between these two states. Although the two-state model of polypeptides has a lot of support, especially revealed in the Ramachandran plots, showing that peptide backbone has two well separated states corresponding to the ␣-helix and the ␤-sheet (the high-temperature denatured coil being the random mixing between the two), from the fieldtheoretical point of view, it is desirable to have a continuum model of chain conformation, as a function of interaction pot...

Protein structure determination by classical x-ray crystallography requires three-dimensional crystals that are difficult to obtain for most proteins and especially for membrane proteins. An alternative is to grow two-dimensional (2D) crystals by adsorbing proteins to ligand-lipid monolayers at the surface of water. This confined geometry requires only small amounts of material and offers numerous advantages: self-assembly and ordering over micrometer scales is easier to obtain in two dimensions; although fully hydrated, the crystals are sufficiently rigid to be investigated by various techniques, such as electron crystallography or micromechanical measurements. Here we report structural studies, using grazing incidence synchrotron x-ray diffraction, of three different 2D protein crystals at the air-water interface, namely streptavidine, annexin V, and the transcription factor HupR. Using a set-up of high angular resolution, we observe narrow Bragg reflections showing long-range crystalline order in two dimensions. In the case of streptavidin the angular range of the observed diffraction corresponds to a resolution of 10 A in plane and 14 A normal to the plane. We show that this approach is complementary to electron crystallography but without the need for transfer of the monolayer onto a grid. Moreover, as the 2D crystals are accessible from the buffer solution, the formation and structure of protein complexes can be investigated in situ.

The quasi-equilibrium evolution of the helical fraction occurring in a biopolymer network (gelatin gel) under an applied stress has been investigated by observing modulation in its optical activity. Its variation with the imposed chain extension is distinctly nonmonotonic and corresponds to the transition of initially coiled strands to induced left-handed helices. The experimental results are in qualitative agreement with theoretical predictions of helices induced on chain extension. This new effect of mechanically stimulated helix-coil transition has been studied further as a function of the elastic properties of the polymer network: crosslink density and network aging.