If it doesn't inhibit your enzyme ), try 0.05 to 0.10% sodium azide.
But you have to test first its effect on your enzyme; some enzymes drop dead
immediately upon exposure to azide. Your membrane extract probably contains
the enzymes responsible for the oxidative phosphorylation.
Cheers,
Achim
Chaitanya Athale <chaitanya.athale at ibc.unibe.ch> wrote in message
news:008601bf0bf5$1c8e8ae0$0b4b5c82 at erni8.unibe.ch...
> This is a multi-part message in MIME format.
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>> Hi everybody. I have a problem: I am trying to set up a typical NADH
coupled assay to measure Pyrvate formation (using LDHase). One of the
proteins of my system is a transmembrane protein, and I have to use it as a
membrane extract. The problem is that in the membrane there seems to exist a
second protein which interferes with the coupled assay, because it consumes
NADH by a different and unknown pathway (I have not found yet the substrate
being reduced). The solution would be very simple if I can find any general
irreversible inhibitor for proteins which use NADH.
> Does anybody know any such inhibitor??
>> Thank you very much for any information.
>>> --Boundary_(ID_aFjp0A6VPEaopwbyV6r8mA)
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> <META content='"MSHTML 4.72.3110.7"' name=GENERATOR>
> </HEAD>
> <BODY bgColor=#ffffff>
> <DIV><FONT color=#000000 face=Arial size=2> <FONT
> color=#000000>Hi everybody. I have a problem: I am trying to
set up
> a typical NADH coupled assay to measure Pyrvate formation (using LDHase).
One of
> the proteins of my system is a transmembrane protein, and I have to use it
as a
> membrane extract. The problem is that in the membrane there seems to exist
a
> second protein which interferes with the coupled assay, because it
consumes NADH
> by a different and unknown pathway (I have not found yet the substrate
being
> reduced). The solution would be very simple if I can find any general
> irreversible inhibitor for proteins which use NADH.</FONT></FONT></DIV>
> <DIV><FONT color=#000000 face=Arial size=2><FONT
> color=#000000> Does anybody know any such
> inhibitor??</FONT></FONT></DIV>
> <DIV><FONT color=#000000 face=Arial size=2><FONT
> color=#000000></FONT></FONT> </DIV>
> <DIV><FONT color=#000000 face=Arial size=2><FONT
> color=#000000> Thank you very much for any
> information.</FONT></FONT></DIV>
> <DIV><FONT color=#000000 face=Arial size=2><FONT
> color=#000000></FONT></FONT> </DIV></BODY></HTML>
>> --Boundary_(ID_aFjp0A6VPEaopwbyV6r8mA)--
