What would happen by replacing ATP with equal amounts of ATP containing radiolab

Question

What would happen by replacing ATP with equal amounts of ATP containing radiolabeled phosphorus at the X marked on the graph? Are enzyme kinetics such as Km, Vmax, Kcat, etc. are affected? What other conclusions can be made from all this information? Good reasoning will be very much appreciated!

I mix together Glucose, ATP and purified Hexokinase and then measure the amount of Glc-6-PO that is formed over time. My data is shown in the graph. At- the point marked "X", I instantaneously replace some of the ATP still present in the reaction with an equal amount of ATP containing radiolabeled phosphorus X- ? Time

Explanation / Answer

The graph shows a hyperbolic curve. It means that the given enzyme follow Michaelis-Menten kinetics. Hexokinase catalyzes the following reaction.

Glucose + ATP ---------> Glucose-6-phosphate + ADP

It is an irreversible reaction. So, if we replace cold ATP with radioactive ATP at point X where the reaction rate (Vmax) is maximum, it does not affect any of the reaction kinetics such as Km, Vmax or Kcat (As long as the total ATP concentration remains constant). The enzyme does not differentiate between unlabelled and radiolabelled ATP. Both serve as enzyme substrates with equal affinity.

Since the concentration or the nature of reactants did not change, reaction kinetics also do not change.

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