Evaluation of mixed-salt effects on thermodynamic and kinetic parameters of RNA polymerase-promoter DNA complexes in terms of equivalent salt concentrations. General applicability to DNA complexes.
Facile evaluation of mixed-salt effect on the strongly salt-dependent thermodynamic and kinetic parameters of protein-DNA complexes is of importance for relevant biochemical and biophysical studies. In pursuit of this aim, binding isotherms for open transcription complex (RPo) of Escherichia coli RNA polymerase (R) at lambdaP(R) promoter DNA (P) were determined as a function of salt concentration in pure NaCl and Tris/HCl solutions, and as a function of [NaCl] in the presence of fixed concentrations of MgCl(2) and Tris/HCl. A concept of equivalent salt concentrations, i.e. concentrations at which the binding equilibrium constant is the same, was introduced and applied for prediction of binding isotherms in mixed salt solutions. Full coincidence between the experimental and predicted isotherms indicated that individual contributions of salts to the global salt-effect are additive in a broad range of salt concentrations. A generalized formula for calculation of salt equivalents characteristic for any of the thermodynamic or kinetic parameters of a complex (e.g., free energy, binding equilibrium and association/dissociation kinetic rate constants) is presented and its applicability to a number of protein-DNA complexes and dsDNA melting demonstrated using authors' own and literature data.
Łoziński
Tomasz
T
Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warszawa, Poland.
Wierzchowski
Kazimierz L
KL
D010582:Bacteriophage lambda / Q000235:genetics; D004247:DNA / Q000378:metabolism; D004279:DNA, Viral / Q000187:drug effects / Q000235:genetics; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D007700:Kinetics; D015636:Magnesium Chloride / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000187:drug effects; D012492:Salts / Q000494:pharmacology*; D012965:Sodium Chloride / Q000494:pharmacology*; D013816:Thermodynamics
D004279:DNA, Viral; D012492:Salts; D015636:Magnesium Chloride; D012965:Sodium Chloride; D004247:DNA; D012321:DNA-Directed RNA Polymerases