How to accurately do Protein/Enzyme Assay Calculations?

Question

This is for my Biochemistry of Macromolecules Lab.....
For determining enzyme concentration (umole/min) for M1, P1, P2, and S2 using the fact that 9OD=1 umole, does these calculations seem reasonable.
The OD's are: M1=.888, P1=.352, P2=.473, and S2= .034
I divided the OD's by nine to yield the umole units and then multiplied by the dilution factor 50 (20:1000) and .020 ml (amount of enzyme used)
To calculate the total units I multiplied the enzyme units by the volume in ml's from the protein assay from the previous week (it is total volume of the M1-S2).
To calculate the specific activity I divided the enzyme concentration by the protein concentration.
To calculate the purification factor, I divided the final specific activity by the initial specific activity and for the % yield of protein and enzyme I simply divided the final total units by the initial total units and multiplied by 100.......
HAS ANYONE DONE THIS BEFORE.....DOES THE CALCULATIONS SOUND RIGHT?????

Answer 1

It is good that you ask for someone to double check your work instead of asking them to do it for you.
We don't know the specifics of the experiment so it is rather difficult to say... Do you take time into account? If the assay time is 1min then you don't need to, since you are expressing in something per min.

I guess you get 9OD=1umole from a standard curve, so yes you correctly divided the OD values to get the # of umoles. From then on I really can't say for the dilutions if it is right since I don't know the experimental procedure.

Specific activity would be enzyme units/volume divided by protein/volume or total units/total protein
Purification factor sounds OK
% yield protein,enzyme should be (final protein,enzyme/initial protein,enzyme) *100

Answer 2

Enzyme Activity Assay Calculations

Answer 3

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