Effects of monovalent anions on a temperature-dependent heat capacity change for Escherichia coli SSB tetramer binding to single-stranded DNA

We have previously shown that the linkage of temperature-dependent protonation and DNA base unstacking equilibria contribute significantly to both the negative enthalpy change (Delta H-obs) and the negative heat capacity change (Delta C-p,C-obs) for Escherichia coli SSB homotetramer binding to single-stranded (ss) DNA. Using isothermal titration calorimetry we have now examined Delta H-obs, over a much wider temperature range (5-60 degrees C) and as a function of monovalent salt concentration and type for SSB binding to (dT)(70) under solution conditions that favor the fully wrapped (SSB)65 complex (monovalent salt concentration >= 0.20 M). Over this wider temperature range we observe a strongly temperature-dependent Delta C-p,C-obs. The Delta H-obs decreases as temperature increases from 5 to 35 degrees C (Delta C-p,C-obs < 0) but then increases at higher temperatures up to 60 degrees C (Delta C-p,C-obs > 0). Both salt concentration and anion type have large effects on Delta H-obs and Delta C-p,C-obs. These observations can be explained by a model in which SSB protein can undergo a temperature- and salt-dependent conformational transition (below 35 degrees C), the midpoint of which shifts to higher temperature (above 35 degrees C) for SSB bound to ssDNA. Anions bind weakly to free SSB, with the preference Br- > Cl- > F-, and these anions are then released upon binding ssDNA, affecting both Delta H-obs and Delta C-p,C-obs. We conclude that the experimentally measured values of Delta C-p,C-obs for SSB binding to ssDNA cannot be explained solely on the basis of changes in accessible surface area (ASA) upon complex formation but rather result from a series of temperature-dependent equilibria (ion binding, protonation, and protein conformational changes) that are coupled to the SSB-ssDNA binding equilibrium. This is also likely true for many other protein-nucleic acid interactions.