In article <305vv0$754 at news.iastate.edu> Bipin K Dalmia,
bipin at iastate.edu writes:
I found this posted on bionet.molbio.methds-reagnts and thought it more
properly belongs on bionet.molbio.proteins:
In article <305vv0$754 at news.iastate.edu> Bipin K Dalmia,
bipin at iastate.edu writes:
>what is the best way (quickest) to adjust the pH of a protein solution?
>dialysis takes too long and desalting/buffer exchange columns are out
>'cause i have 100 mL of solution and it would take a 10-story high
>column. directly adding acid/base would cause extreme local pH values
>and denature the protein.
>>i have to do this prior to running an ion-exchange column so i can't
>increase the ionic-strength too much either.
I wonder about the answer I posted:
>First, the disclaimer: I never was much of a protein chemist, and it
has been 20 years >since I even pretended to know what I was doing with
them. I highly recommend >checking with someone really knowledgeable
before taking any of _my_ suggestions:
>>1) Dialysis need not take very long. Pencil-wide dialysis tubing which
is stirred along >with the exterior buffer would be 95% equilibrated in
half an hour. Granted 100ml >would require a lot of this thin tubing.
But 3 inch tubing for 60-90 minutes might> >work OK as long as the bag
and its contents are stirred vigorously.
>>2) Acetone precipitation followed by dissolving in the buffer of choice
should work as >long as the protein can be easily redissolve.
>>3) Ammonium sulfate precipitation, with the same caveat.
>>4) TCA precipitation, same caveat.
>>5) You could dilute your 100ml into a liter or two of what you want for
the ion >exchange chromatography and then run the ion exchange. Ion
exchange chromatography >can concencentrate the protein as long as you do
not exceed the capacity of the resin.
>>Those are my ideas for what they are worth. (Every penny you paid.) A
real protein >chemist might want to know more details about the
properties of your protein before >making suggestions, but I am a fool.
Perhaps you should call technical support (for >industrial applications)
of Pharmacia.
Any comments? Please cross post to bionet.molbio.methds-reagnts
Jim Owens
