[LapSum]

Hi all,
we are doing an enzymatic reaction, in which acetyl co-A is used by an emzyme to acetylate a proteins subtrate at pH 6-7 in room temperature.
The reaction release free co-A which is detected by a dye as a readout.

The problem is, the acetyl-coA itself is not 100% stable and it breaks down and attach the aetyl group to the protein subtract without the help of any enzyme (the rate of this reaction is very low tho)
We do want to completely eliminate this background noise.

I wonder if there is a way to either
1. make the acetyl coA more stable (in certain pH? the presence of certain chemicals to inhibit this?)
2. stop the broke down reaction acetyl group from attacking our protein without the enzyme.

I am a biologist, am certainly not good in chemisty. Please help. Thanks so much!

[Babcock_Hall]

I would find a bioorganic textbook, and look for thioester stability as a function of pH.  A model thioester would be almost as good as acetyl CoA itself.  The old ones (Bruice and Benkovic; Jencks) might be as good or better than the new ones for this sort of information.  I think you have two side reactions:
a) hydrolysis of acetyl CoA (other than pH and temperature, there is not much you can do)
b) nonenzymatic reaction of acetyl CoA with your protein.

You may not be able to stop b).  However if you do a plot of rate of acetylation versus [enzyme], the y-intercept should be the rate of nonenzymatic acetylation.  That's all I can think of...