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DOI

Journal: Nature

Experimental demonstration of a single-molecule electric motor

Heather L. Tierney, Colin J. Murphy, April D. Jewell, Ashleigh E. Baber, Erin V. Iski, Harout Y. Khodaverdian, Allister F. McGuire, Nikolai Klebanov, and E. Charles H. Sykes



For molecules to be used as components in molecular machines, methods that couple individual molecules to external energy sources and that selectively excite motion in a given direction are required. Significant progress has been made in the construction of molecular motors powered by light and by chemical reactions, but electrically driven motors have not yet been built, despite several theoretical proposals for such motors. Here we report that a butyl methyl sulphide molecule adsorbed on a copper surface can be operated as a single-molecule electric motor. Electrons from a scanning tunnelling microscope are used to drive the directional motion of the molecule in a two-terminal setup. Moreover, the temperature and electron flux can be adjusted to allow each rotational event to be monitored at the molecular scale in real time. The direction and rate of the rotation are related to the chiralities of both the molecule and the tip of the microscope (which serves as the electrode), illustrating the importance of the symmetry of the metal contacts in atomic-scale electrical devices.
 DOI
Journal: Nature Nanotechnology

Long-Lived States to Monitor Protein Unfolding by Proton NMR

1. Aurélien Bornet, 
2. Puneet Ahuja, 
3. Riddhiman Sarkar, 
4. Laetitia Fernandes, 
5. Sonia Hadji, 
6. Shirley Y. Lee, 
7. Aydin Haririnia,
8.  David Fushman, 
9. Geoffrey Bodenhausen, and 
10. Paul R. Vasos

1. The relaxation of long-lived states (LLS) corresponds to the slow return to statistical thermal equilibrium between symmetric and antisymmetric proton spin states. This process is remarkably sensitive to the presence of external spins and can be used to obtain information about partial unfolding of proteins. We detected the appearance of a destabilized conformer of ubiquitin when urea is added to the protein in its native state. This conformer shows increased mobility in the C-terminus, which significantly extends the lifetimes of proton LLS magnetisation in Ser-65. These changes could not be detected by conventional measurements of T₁ and T₂ relaxation times of protons, and would hardly be sensed by carbon-13 or nitrogen-15 relaxation measurements. Conformers with similar dynamic and structural features, as revealed by LLS relaxation times, could be observed, in the absence of urea, in two ubiquitin mutants, L67S and L69S.

1. DOI

1. Journal: ChemPhysChem

Optical trapping for the characterization of amyloid-beta aggregation kinetics

Anthony J. Veloso, Hiroyuki Yoshikawa, Xin R. Cheng, Eiichi Tamiya, and Kagan Kerman

Alzheimer's disease (AD) is marked by the accumulation of neuronal plaques from insoluble amyloid-beta (Aβ) peptides. Growing evidence for the role of Aβ oligomers in neuronal cell cytotoxicity and pathogenesis has prompted the development of novel techniques to better understand the early stages of aggregation. Near infrared (NIR) optical trapping was applied to characterize the early stages of Aβ aggregation in the presence of a β-sheet intercalating dye, Congo Red (CR), as the fluorescent marker. The integration of fluorescence analysis with NIR optical trapping has provided a new outlook into the first two hours of Aβ aggregation.

DOI

Journal: Analyst

Direct Quantification of the Attempt Frequency Determining the Mechanical Unfolding of Ubiquitin Protein

Ionel Popa, Julio M. Fernández, and Sergi Garcia-Manyes

Understanding protein dynamics requires a comprehensive knowledge of the underlying potential energy surface that governs the motion of each individual protein molecule. Single molecule mechanical studies have provided the unprecedented opportunity to study the individual unfolding pathways along a well defined coordinate, the end-to-end length of the protein. In these experiments, unfolding requires surmounting an energy barrier that separates the native from the extended state. The calculation of the absolute value of the barrier height has traditionally relied on the assumption of an attempt frequency, υ^‡. Here we used single molecule force-clamp spectroscopy to directly determine the value of υ^‡ for mechanical unfolding by measuring the unfolding rate of the small protein ubiquitin at varying temperatures. Our experiments demonstrate a significant effect of the temperature on the mechanical rate of unfolding. By extrapolating the unfolding rate in the absence of force for different temperatures, varying within the range spanning from 5 to 45 °C, we measured a value for the activation barrier of ΔG^‡ = 71 ± 5 kJ/mol and an exponential prefactor υ^‡ ∼4 × 10⁹ s⁻¹. Although the measured prefactor value is 3 orders of magnitude smaller than the value predicted by the transition state theory (∼6 × 10¹² s⁻¹), it is 400-fold higher than that encountered in analogous experiments studying the effect of temperature on the reactivity of a protein-embedded disulfide bond (∼10⁷ M⁻¹ s⁻¹). This approach will allow quantitative characterization of the complete energy landscape of a folding polypeptide from highly extended states, of capital importance for proteins with elastic function.

DOI

Journal: The Journal of Biological Chemistry

Myosin Va and myosin VI coordinate their steps while engaged in an in vitro tug of war during cargo transport

M. Yusuf Ali, Guy G. Kennedy, Daniel Safer, Kathleen M. Trybus, H. Lee Sweeney, and David M. Warshawa

Myosin Va (myoV) and myosin VI (myoVI) are processive molecular motors that transport cargo in opposite directions on actin tracks. Because these motors may bind to the same cargo in vivo, we developed an in vitro “tug of war” to characterize the stepping dynamics of single quantum-dot-labeled myoV and myoVI motors linked to a common cargo. MyoV dominates its myoVI partner 79% of the time. Regardless of which motor wins, its stepping rate slows due to the resistive load of the losing motor (myoV, 2.1 pN; myoVI, 1.4 pN). Interestingly, the losing motor steps backward in synchrony with the winning motor. With ADP present, myoVI acts as an anchor to prevent myoV from stepping forward. This model system emphasizes the physical communication between opposing motors bound to a common cargo and highlights the potential for modulating this interaction by changes in the cell’s ionic milieu.

DOI

Journal: Proceedings of the National Academy of Sciences

Freely orbiting magnetic tweezers to directly monitor changes in the twist of nucleic acids

Jan Lipfert, matthew Wiggin, Jacob W.J. Kerssemakers, Francesco Pedaci, and Nynke H. Dekker


The double-stranded nature of DNA links its replication, transcription and repair to rotational motion and torsional strain. Magnetic tweezers (MT) are a powerful single-molecule technique to apply both forces and torques to individual DNA or RNA molecules. However, conventional MT do not track rotational motion directly and constrain the free rotation of the nucleic acid tether. Here we present freely orbiting MT (FOMT) that allow the measurement of equilibrium fluctuations and changes in the twist of tethered nucleic acid molecules. Using a precisely aligned vertically oriented magnetic field, FOMT enable tracking of the rotation angle from straight forward (x,y)-position tracking and permits the application of calibrated stretching forces, without biasing the tether's free rotation. We utilize FOMT to measure the force-dependent torsional stiffness of DNA from equilibrium rotational fluctuations and to follow the assembly of recombination protein A filaments on DNA.


DOI


Journal: Nature Communications