phospho to total protein

[scholar]

Why is the ratio mainly used as the standard to look at protein activation? Is n't the total amount of phospho protein more important regardless of how much total there is?

[admin]

I guess the ratio looks at the percentage of phosphorylated protein. The problem is with phosphatases is you will rarely get the total phosphorylated protein.

Even with total protein (with proteases and inaccuracies), and the measured phosphorylation percentage (with phosphatase, detection problems and inaccuracies) this is only a good estimate with western blotting.

The most accurate is to use radiolabelling or similar non-radiolabelled signal however a lot of research is focusing on western blot techniques

[scholar]

I understand that the ratio gives you a percentage of activated protein, and that phosphatases will affect the absolute amount of phosphoprotein at a given time. However, phosphatase effects are still present in the ratio (since you are just measuring the same thing but dividing by total proteins). To me, it seems like you are getting the same amount of information, unless the total levels of phosphoprotein changes upon your perturbation.

I am still not satisfied explanations given thus far, aside from "it is the standard in the field". I mean it is really a simple question.

Scholar