Klenow and Klentaq Pol I DNA polymerases from E. coli and Thermus aquaticus both bind DNA with high affinity and large heat capacity changes, resulting in a significant variation in the binding enthalpy versus temperature.  Functionally, both polymerases are catalytically inactive until the temperature where the binding enthalpy crosses zero (TH) and becomes favorable.  These data indicate that a negative free energy of DNA binding alone is not sufficient to drive catalysis, and that a negative enthalpy of initial binding (?H) is required for nucleotide incorporation activity.  TH for Klentaq is 15 degrees higher than for Klenow, indicating that there is not a universal polymerase activation temperature.  Analysis of the temperature dependence of the catalytic activity of the polymerases using Eyring theory yields patterns for ?G�, ?H�, and T?S� analogous to those commonly observed for direct DNA binding, thus also yielding an apparent ?Cp� for forming the activated complex.

Enthalpic on/off switch linking DNA binding and nucleotide incorporation activity in Pol I DNA polymerases

245th National Meeting (2013)

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