We obtain measurements of the elastic energy of short (18-30 bp) molecules of ds DNA constrained into a sharply bent conformation, using a thermodynamic method with the DNA in solution. We consider the case where there is one nick in the ds DNA, and find that the system develops a kink at a critical torque τc ≈ 27 pN × nm. In this regime the elastic energy is linear in the end-to-end distance (EED). For smaller torques the DNA is smoothly bent and described by the worm-like-chain energy, which is also approximately linear in the EED, but with a different slope. Thus we access both the high and low elastic energy regimes, and the transition between the two.

We measure the bending energy of double-stranded DNA in the nonlinear (sharply bent) regime. The measurements are obtained from the melting curves of stressed DNA ring molecules. The nonlinear elastic behavior is captured by a single parameter: the critical torque c at which the molecule develops a kink. In this regime, the elastic energy is linear in the kink angle. This phenomenology is the same as for the previously reported case of nicked DNA. For the sequences examined, we find c ¼ 31 pN Â nm. This critical torque corresponds to a characteristic energy scale ð =2Þ c ¼ 12 kT room relevant for molecular biology processes associated with DNA bending.

We address the coupling of mechanics and chemistry in an enzyme through equilibrium experiments where we mechanically deform the enzyme and measure the effect on the chemical reaction it catalyzes. We use the DNA molecular spring method to exert stresses at three different specific locations on the enzyme Guanylate Kinase, and for each case determine the changes in substrates binding affinities and catalytic rate. We find that the enzyme kinetics parameters can be affected separately, depending on where the mechanical stress is applied. For one configuration the applied stress mainly affects the catalytic rate kcat, for another it mainly affects the binding affinity of the substrate GMP. These experiments show that a stress applied by pulling two residues on the surface of the protein generally results in a strain propagating into the structure.

We report experiments where the activity of the enzyme luciferase from Renilla reniformis is controlled through a DNA spring attached to the enzyme. In the wake of previous work on kinases, these results establish that mechanical stress applied through the DNA springs is indeed a general method for the artificial control of enzymes, and for the quantitative study of mechano-chemical coupling in these molecules. We also show proof of concept of the luciferase construct as a sensitive molecular probe, detecting a specific DNA target sequence in an easy, one-step, homogeneous assay, as well as SNP detection without melting curve analysis.

We present experimental measurements of the equilibrium elastic energy of protein-DNA chimeras, for two different sets of attachment points of the DNA "molecular spring" on the surface of the protein. Combining these with measurements of the enzyme's activity under stress and a mechanical model of the system, we determine how the elastic energy is partitioned between the DNA and the protein. The analysis shows that the protein is mechanically stiffer than the DNA spring.