pH-buffer system

Kresten kresten at my-deja.com
Thu Oct 26 07:37:22 EST 2000


Depending on what reaction your enzyme is catalyzing, it might be a
good idea to avoid "reactive" buffers, e.g. good nucleophiles like
Tris. Also, perhaps it is possible to use two different buffer at some
pH values as in the following example:

pH   buffer#
5    1
5.5  1
6    1
6    2
6.5  2
7    3
7    4

etc

If the two activities at eg pH 6 are different then you know that
something is fishy.

Alternatively you could run the reaction in a pH stat, perhaps in the
presence of eg KCl to mask the effect of the titrand.

Your suggestion that you merely mix the buffers may give you a false
feeling of being safe because the interference of the buffers on your
assay may be pH dependent. That is having 50mM tris in a phosphate
buffer at pH 6.5 may be irrelevant but in a carbonate buffer pH 10 it
might be important.

HTH
Kresten

In article <39F81C3E.9C381EE3 at vub.ac.be>,
  Sigrid Van Boxstael <svboxsta at vub.ac.be> wrote:
> Hi,
>
> I'd like to determine the activity of my protein in a whole pH-range
> from 5 to 11.
> I could use different buffers, like MES, CAPS, MOPS, CHES, Tris, ...
> who
> all have a
> buffering-capacity around their pKa.
> If  I would use them all separately, I presume that there might be an
> influence of the
> type of buffer on my activity.
> Does anyone now either it is adivisable to mix the buffers and adjust
> to
> different pH's?
> Does anyone have a proposal for a buffer system or maybey a reference?
>
> Thanks,
> Sigrid
>
>

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Kresten Lindorff Larsen


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