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2005
LSU-HHMI Summer Undergraduate Research Program |
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Chelsea Agar and Vince J. LiCata, Biological Sciences
A Comparison of the effects of bound DNA on the thermostability
of Type I DNA polymerases from Thermus aquaticus and Escherichia
coli
Klenow and Klentaq, the large fragment domains of DNA polymerase
I from Escherichia coli and Thermus aquaticus respectively,
were examined when bound to a 13/20mer DNA segment using circular
dichroism (CD) spectroscopy and differential scanning calorimetry
(DSC). Thermal denaturations of both species, alone and as a
DNA-protein complex, were preformed to directly measure the
effects of bound DNA on melting temperature (Tm), constant pressure
heat capacity (DCp), and enthalpy (DH), and to help elucidate
the basis for the more thermally stable nature of Klentaq versus
its mesophillic homolog, Klenow. Experimental data from both
DSC and CD show essentially no difference in Tm for Klentaq
upon DNA binding, whereas both techniques show a significant
change in Tm between Klenow and the Klenow-DNA complex. It was
determined that bound DNA has a stabilizing effect on Klenow
but no stabilizing effect on the already extremely thermally
stable protein Klentaq.
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