Fluorescence depolarization and temperature dependence of the torsion elastic constant of linear phi 29...

The torsion elastic constant alpha of linear phi 29 DNA has been determined as a function of temperature from 10 to 78 degrees C by studying the decay of the fluorescence polarization anisotropy (FPA) of intercalated ethidium dye. The time-dependent FPA was measured by using a picosecond dye laser for excitation and time-correlated single photon counting detection. Over the region 10-74 degrees C, alpha was effectively constant within experimental error, varying from (3.5 +/- 0.4) X 10(-12) dyn cm at 10 degrees C to (3.7 +/- 0.3) X 10(-12) dyn cm at 74 degrees C. At 78 degrees C, which is just above the melting temperature Tm = 76 degrees C, alpha decreased to (3.3 +/- 0.3) X 10(-12) dyn cm, and at 90 degrees C, where the DNA is completely denatured, both the fluorescence lifetime and the decay time of the FPA are characteristic of unbound ethidium bromide. The weak temperature dependence of alpha implies that DNA torsional deformations do not occur primarily at sites of high enthalpy perturbed structures such as open base pairs.