Influence of electric dichroism on the temperature-jump relaxation study of proflavine-DNA complexes

The temperature-jump relaxation kinetics of proflavine-DNA complexes has been reinvestigated with a standard apparatus equipped for absorption detection of plane-polarized light in order to discriminate between chemical relaxation and transient orientation effects. Under low ionic strength conditions (0.015 M Na+), these effects may represent the major contribution to the signal when the T-jump apparatus is used without a polarizer. They have been improperly assigned to chemical relaxation in previous work. The actual relaxation times are smaller than 30 musec at 10 degrees C. Under medium ionic strength conditions (0.2 M Na+) it is shown that: (i) the "instantaneous" change of transmission reported in earlier work (Li, H. J., and Crothers, D. M. (1969), J. Mol. Biol. 39, 461-477; Schmechel, D. E. V., and Crothers, D. M. (1971), Biopolymers 10, 465-480) is due to orientation effects; (ii) an intermediate exists whose absorption spectrum resembles somewhat that of proflavine aggregates on a linear polyanion; (iii) the rate constants for outside binding may be significantly larger than previously reported. The new kinetic data are consistent with a modified mechanism derived from equilibrium studies (Ramstein, J., Hogrel, J. F., Dourlent, M., Leng, M., an Hélène, C. 1973), in Dynamic Aspects of Conformation Changes in Biological Macromolecules, Sadron, C., Ed., Dordrecht, Holland, Reidel Publisher, pp 333-347; Dourlent, M., and Hogrel, J. F. (1976), Biopolymers (in press)), but, however, do not unambiguously prove it. From these studies, it is inferred that orientation effects can perturb relaxation data on systems containing linear polymers in many cases.