Finding a suitable denaturing agent

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cindyfunnybaby
cindyfunnybaby's picture
Finding a suitable denaturing agent

Hi, everyone

I got some problems in denaturing the endogenous collagenase of planarian extract. I had try to use 8 M of urea and complete protease inhibitor for extraction, but I still cannot inhibit the collagenase activity.
what another denaturing agent that I can used to solve this problem? Does guanidine - HCl is a good choice?

thank you v.much~

kjosephs
kjosephs's picture
cindyfunnybaby wrote:Hi,

cindyfunnybaby wrote:

Hi, everyone I got some problems in denaturing the endogenous collagenase of planarian extract. I had try to use 8 M of urea and complete protease inhibitor for extraction, but I still cannot inhibit the collagenase activity. what another denaturing agent that I can used to solve this problem? Does guanidine - HCl is a good choice? thank you v.much~

GuHCl at 6M is also worth trying.  You can also try  heating your sample in the 8M Urea or 6 M GuHCl.  Does collagenase have disulfide bonds?  Are you adding reductant, DTT or BME?
I have not used GuHCl at higher than 6M.  Anyone else?

R Bishop
R Bishop's picture
Im pretty sure GuHCL will

Im pretty sure GuHCL will denature anything at 4M.  Will try to find a reference.