ATPase assay

Sam bovis drb13 at cornell.edu
Fri Nov 22 20:35:40 EST 1996


> I isolated and expressed (in E. coli) a couple new proteins. I wonder
how can I an
> alysis the potential ATPase activity of them, in another word, how do I
start? Any 
> suggestion is welcomed. Thanks.
> 
>                                         Ke Wu
Incubate your sample with ATP, stop the reaction with acid, and analyze
for phosphorus using any standard colorometric assay.  

Or, if you can work at near-neutral pH,  do a coupled assay with pyruvate
kinase, PEP, NADH and lactate dehydrogenase. When you add ATP,  the
appearance of ADP will cause NADH oxidation. Following the rate of derease
at 340 nm gives an instant reading of the ATPase activity.

Both of these can be found in most literature dealing with isolation or
characterization of ATPases.



More information about the Proteins mailing list