Nucleic Acids Research, 2003, Vol. 31, No. 19 5590-5597
© 2003 Oxford University Press
Thermodynamics of the binding of Thermus aquaticus DNA polymerase to primed-template DNA
Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA
*To whom correspondence should be addressed. Tel: +1 225 578 5233; Fax: +1 225 578 2597; Email: licata{at}lsu.edu
DNA binding of the Type 1 DNA polymerase from Thermus aquaticus (Taq polymerase) and its Klentaq large fragment domain have been studied as a function of temperature. Equilibrium binding assays were performed from 5 to 70°C using a fluorescence anisotropy assay and from 10 to 60°C using isothermal titration calorimetry. In contrast to the usual behavior of thermophilic proteins at low temperatures, Taq and Klentaq bind DNA with high affinity at temperatures down to 5°C. The affinity is maximal at 40–50°C. The 
H and S of binding are highly temperature dependent, and the Cp of binding is –0.7 to –0.8 kcal/mol K, for both Taq and Klentaq, with good agreement between van’t Hoff and calorimetric values. Such a thermodynamic profile, however, is generally associated with sequence-specific DNA binding and not non- specific binding. Circular dichroism spectra show conformational rearrangements of both the DNA and the protein upon binding. The high Cp of Taq/Klentaq DNA binding may be correlated with structure-specific binding in analogy to sequence- specific binding, or may be a general characteristic of proteins that primarily bind non-specifically to DNA. The low temperature DNA binding of Taq/Klentaq is suggested to be a general characteristic of thermophilic DNA binding proteins.
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