T4 PNK

peter via methods%40net.bio.net (by peter.ianakiev from gmail.com)
Mon Jun 30 15:33:16 EST 2008


On Jun 30, 10:49 am, "Tim Fitzwater" <tfitzwa... from somalogic.com> wrote:
> Lillehaug, J. R. and K. Kleppe (1975). "Kinetics and specificity of T4
> polynucleotide kinase." Biochemistry 14(6): 1221-5.
>
>                 The kinetics of T4 polynucleotide kinase has been
> investigated at pH 8.0 and 37 degrees. Double reciprocal plots of
> initial rates vs. substrate concentrations as well as product inhibition
> studies have indicated that the enzyme reacts according to the ordered
> sequential mechanism shown in eq 2 in the text for phosphorylation of a
> DNA molecule. Based on this mechanism the rate equation for the overall
> reaction was deduced and the various kinetic constants estimated. Hill
> plots indicated little or no interaction between active sites in the
> enzyme. The apparent Michaelis constants and V-max were determined at a
> fixed ATP concentration, 66 muM, for a number of different substrates
> varying in chain length, base composition, and nature of the sugar, and
> a wide variation was found. For the nucleoside 3'-monophosphates tested
> both the apparent Michaelis constant and V-max values were from
> approximately 2 to 5 times larger than for the corresponding
> oligonucleotide. The following orders were obtained with regard to
> apparent Michaelis constants and V-max for the nucleoside
> 3'-monophosphates investigated: Michaelis constant, rGP greater than rUp
> greater than rCp greater than rAp greater than dTp; V-max, rGp greater
> than rCp greater than rAp greater than dTp greater than rUp. Somewhat
> similar results were also obtained with the deoxyoligonucleotides
> tested.

Whats your point? My question was if T4PNK can use dATP instead of
rATP.
Thanks,
Peter


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