Light-powered molecular machines are conjectured to be essential constituents of future nanoscale devices. As a model for such systems, we have synthesized a polymer of bistable photosensitive azobenzenes. Individual polymers were investigated by single-molecule force spectroscopy in combination with optical excitation in total internal reflection. We were able to optically lengthen and contract individual polymers by switching the azo groups between their trans and cis configurations. The polymer was found to contract against an external force acting along the polymer backbone, thus delivering mechanical work. As a proof of principle, the polymer was operated in a periodic mode, demonstrating for the first time optomechanical energy conversion in a single-molecule device.

Single-molecule Förster resonance energy transfer (smFRET) is increasingly being used to determine distances, structures, and dynamics of biomolecules in vitro and in vivo. However, generalized protocols and FRET standards to ensure the reproducibility and accuracy of measurements of FRET efficiencies are currently lacking. Here we report the results of a comparative blind study in which 20 labs determined the FRET efficiencies (E) of several dye-labeled DNA duplexes. Using a unified, straightforward method, we obtained FRET efficiencies with s.d. between ±0.02 and ±0.05. We suggest experimental and computational procedures for converting FRET efficiencies into accurate distances, and discuss potential uncertainties in the experiment and the modeling. Our quantitative assessment of the reproducibility of intensity-based smFRET measurements and a unified correction procedure represents an important step toward the validation of distance networks, with the ultimate aim of achieving reliable structural models of biomolecular systems by smFRET-based hybrid methods.

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AFM based single molecule force spectroscopy was used for the investigation of single polyelectrolyte chains. Namely, the elasticity of polyvinylamine chains and their desorption from solid surfaces was studied as a function of the polymer's charge density and electrolyte concentration. Experimental force−distance profiles were fitted by the wormlike chain model, including elastic contributions arising from the stretching of bond angles and covalent bonds. It was found that, under the high stretching forces which can be applied in the AFM experiments, the bending rigidity of polyelectrolyte chains (as described by the persistence length) is significantly lower than predicted by Odijk−Skolnick−Fixman (OSF) theory. Furthermore, the desorption force of single physisorbed polymer chains from negatively charged silica surfaces was determined. In addition to the electrostatic interaction between polymer and substrate, which depends linearly on the Debye screening length and the polymer's line charge density, a constant nonelectrostatic contribution to the desorption force was observed.

The molecular chaperone heat-shock protein 90 (Hsp90) is one of the most abundant proteins in unstressed eukaryotic cells. Its function is dependent on an exceptionally slow ATPase reaction that involves large conformational changes. To observe these conformational changes and to understand their interplay with the ATPase function, we developed a single-molecule assay that allows examination of yeast Hsp90 dimers in real time under various nucleotide conditions. We detected conformational fluctuations between open and closed states on timescales much faster than the rate of ATP hydrolysis. The compiled distributions of dwell times allow us to assign all rate constants to a minimal kinetic model for the conformational changes of Hsp90 and to delineate the influence of ATP hydrolysis. Unexpectedly, in this model ATP lowers two energy barriers almost symmetrically, such that little directionality is introduced. Instead, stochastic, thermal fluctuations of Hsp90 are the dominating processes.

During the last half century, identification of an ideal (predominantly entropic) protein elastomer was generally thought to require that the ideal protein elastomer be a random chain network. Here, we report two new sets of data and review previous data. The first set of new data utilizes atomic force microscopy to report single-chain force-extension curves for (GVGVP) 251 and (GVGIP) 260 , and provides evidence for single-chain ideal elasticity. The second class of new data provides a direct contrast between lowfrequency sound absorption (0.1-10 kHz) exhibited by random-chain network elastomers and by elastin protein-based polymers.Earlier composition, dielectric relaxation (1-1000 MHz), thermoelasticity, molecular mechanics and dynamics calculations and thermodynamic and statistical mechanical analyses are presented, that combine with the new data to contrast with random-chain network rubbers and to detail the presence of regular non-random structural elements of the elastin-based systems that lose entropic elastomeric force upon thermal denaturation.The data and analyses affirm an earlier contrary argument that components of elastin, the elastic protein of the mammalian elastic fibre, and purified elastin fibre itself contain dynamic, non-random, regularly repeating structures that exhibit dominantly entropic elasticity by means of a damping of internal chain dynamics on extension.

Experimental single-molecule stretching curves for three backbone architectures (single-stranded DNA, various types of peptides, polyvinylamine) are quantitatively compared with corresponding quantum-chemical (zero-temperature) ab-initio calculations in the high-force range of up to two nanonewtons. For high forces, quantitative agreement is obtained with the contour length of the polymers as the only fitting parameter. For smaller forces, the effects of chain fluctuations are accounted for by using recent theoretical results for the stretching response of a freely-rotating-chain model.

The hydrophobic effect, i.e., the poor solvation of nonpolar parts of molecules, plays a key role in protein folding and more generally for molecular self-assembly and aggregation in aqueous media. The perturbation of the water structure accounts for many aspects of protein hydrophobicity. However, to what extent the dispersion interaction between molecular entities themselves contributes has remained unclear. This is so because in peptide folding interactions and structural changes occur on all length scales and make disentangling various contributions impossible. We address this issue both experimentally and theoretically by looking at the force necessary to peel a mildly hydrophobic single peptide molecule from a flat hydrophobic diamond surface in the presence of water. This setup avoids problems caused by bubble adsorption, cavitation, and slow equilibration that complicate the much-studied geometry with two macroscopic surfaces. Using atomic-force spectroscopy, we determine the mean desorption force of a single spider-silk peptide chain as F ‫؍‬ 58 ؎ 8 pN, which corresponds to a desorption free energy of Ϸ5 kBT per amino acid. Our all-atomistic molecular dynamics simulation including explicit water correspondingly yields the desorption force F ‫؍‬ 54 ؎ 15 pN. This observation demonstrates that standard nonpolarizable force fields used in classical simulations are capable of resolving the fine details of the hydrophobic attraction of peptides. The analysis of the involved energetics shows that water-structure effects and dispersive interactions give contributions of comparable magnitude that largely cancel out. It follows that the correct modeling of peptide hydrophobicity must take the intimate coupling of solvation and dispersive effects into account.atomic-force microscopy ͉ hydrophobic effect ͉ molecular dynamics simulation ͉ single molecules ͉ protein adsorption F or scientists working with biological or soft matter systems, understanding what holds the world together largely means unraveling the mechanism behind the so-called hydrophobic effect. The term hydrophobic attraction (HA) was initially introduced to describe the attraction between small nonpolar molecules such as methane in water (1, 2). It is nowadays more broadly used to describe forces between all kinds of nonpolar objects in aqueous environments, implying a common mechanism for protein folding, micellization, self-assembly of lipids, oil-water demixing, and thus any supermolecular aggregation in water (3). For predicting protein structures and function the magnitude and nature of the HA acting between peptide segments is a central issue that has not been fully resolved. Much effort was put in force measurements between well defined model systems, for example mica surfaces made hydrophobic or micrometer-sized plastic beads. However, these systems are notoriously plagued by secondary effects, such as bubble adsorption and cavitation effects (4, 5) or compositional rearrangements (6). In simulations of interacting planar plates, similar eff...