We have used constant force axial optical tweezers to understand the subtle effects of sequence variations on the mechanical properties of DNA. Using designed sequences of DNA with nearly identical curvatures, but varied AT content, we have shown the persistence length to be highly dependent on the elasticity of DNA. The persistence length varies by almost thirty percent between sequences containing 61% AT and 45% AT. The biological implications of this can be substantial, as the need to bend DNA is involved in a host of regulatory schemes, ranging from nucleosome positioning to the formation of protein-mediated repressor and enhancer loops.

No abstract

Proteolytic degradation of fibrin, the major structural component in blood clots, is critical both during normal wound healing and in the treatment of ischemic stroke and myocardial infarction. Fibrin-containing clots experience substantial strain due to platelet contraction, fluid shear, and mechanical stress at the wound site. However, little is understood about how mechanical forces may influence fibrin dissolution. We used video microscopy to image strained fibrin clots as they were degraded by plasmin, a major fibrinolytic enzyme. Applied strain causes up to 10-fold reduction in the rate of fibrin degradation. Analysis of our data supports a quantitative model in which the decrease in fibrin proteolysis rates with strain stems from slower transport of plasmin into the clot. We performed fluorescence recovery after photobleaching (FRAP) measurements to further probe the effect of strain on diffusive transport. We find that diffusivity perpendicular to the strain axis decreases exponentially with increasing strain, while diffusivity along the strain axis remains unchanged. Our results suggest that the properties of the fibrin network have evolved to protect mechanically loaded fibrin from degradation, consistent with its function in wound healing. The pronounced effect of strain upon diffusivity within fibrin networks offers a means of tuning the transport of proteins and other soluble factors within fibrin-based biomaterials, potentially addressing a key challenge in engineering complex tissues in vitro. The mechanical and functional properties of DNA arise from its double helical structure. It is now widely accepted that the torsional properties of DNA and DNA supercoiling play an important role in the kinetics of many DNAbinding proteins, but the mechanism underlying this relationship remains unclear. To address this gap in our understanding, we need an instrument that can accurately measure and control torsional stress applied to DNA. We have developed a high-bandwidth electromagnetic trapping system that can generate a uniform magnetic field in the sample region and apply constant torque above 10 2 pN$nm on the samples under study. The octupole magnetic trap is integrated into a microscope-based particle tracking system and can rotate superparamagnetic particles with three degrees of rotational freedom. The large signal bandwidth of the current in the coils can reach above 3kHz at 800uH inductive load and the heat generated by the current is dissipated by an active PID-controlled cooling system to prevent heating biological samples. The magnetic trap is being designed to independently control force and torque, allowing us to confine superparamagnetic particles in a trap with low torsional stiffness that is suitable for torque application and measurement at biologically relevant scales. To directly measure the torsional strain in DNA, we are planning to use superparamagnetic beads coated with metal on one hemisphere. Our magnetic torque tweezers are intended to quantitatively measure the changes of torsion...