Nonspecific interactions of Escherichia coli RNA polymerase with native and denatured DNA: differences in the binding behavior of core and holoenzyme

作者：Pieter L. deHaseth, Timothy M. Lohman, Richard R. Burgess, M. Thomas Record · 发表于：Biochemistry · 年份：1978 · DOI：10.1021/bi00602a006 · 被引用次数：166 · 研究领域：Diffusion and Search Dynamics、Bacterial Genetics and Biotechnology、DNA and Nucleic Acid Chemistry

We have investigated the nonspecific interactions of Escherichia coli RNA polymerase core and holoenzyme with double-stranded (ds) and single-stranded (ss) DNA. Binding constants for these interactions as functions of such solution variables as monovalent and/or divalent cation concentration, temperature, or pH were determined by the method of deHaseth et a. [deHaseth, P.L., Gross, C.A., Burgess, R.R. and Record, M.T. (1977), Biochemistry 16, 4777--4783] from analysis of the elution of the proteins from small columns containing immobilized DNA. This technique, although as yet empirical, has been demonstrated to yield accurate binding constants fot the nonspecific interation of lac repressor with ds DNA. We find that observed binding constants (Kobsd) are extraordinarily sensitive functions of the monovalent cation concentration for the interactions of both core and holoenzyme with ds DNA. In the absence of divalent cations, the derivatives --(d log Kobsd/d log [Na+]) are 11 +/- 2 for the holo--ds DNA interaction and 21 +/- 3 for the core--ds DNA interaction. Consequently, approximately 11 and 21 low-molecular-weight ions are released, iin the thermodynamic sense, in the formation of the holo--ds and core--ds complexes, respectively (Record, M.T., Jr., Lohman, T.M., and deHaseth, P.L. (1976), J. Mol. Biol. 107, 145--158; Record, M.T., Jr., Anderson, C.F., and Lohman, T.M. (1978), Q. Rev. Biophys., in press). Ion release is a thermodynamic driving force for these nonspecific in...