Cleavage by thrombin

Marc Champagne M.Champagne at -nospam-cellbio.duke.edu
Sat Jul 12 08:29:31 EST 1997


In article <33C0EA6E.41C6 at gandalf.psf.sickkids.on.ca>, Randy Willis
<willis at gandalf.psf.sickkids.on.ca> wrote:

> Simon Hoffenberg wrote:
> >We used high thrombin concentration to
> > accelerate the digestion and lost our protein. Any cure for the problem?
> 
> Simon,
> 
> Try using less thrombin and more patience.  We've not experienced too
> many problems with non-specific cleavage for reactions which have
> proceeded for up to 24h.
> 
> Good luck,
> 
> Randall C Willis, Researcher
> Biochemistry Research, Hospital for Sick Children
> 3522-555 University Ave
> Toronto, ON
> M5G 1X8  CANADA
> 
> 416-813-5933 (ph)  -5022 (fax)
> willis at gandalf.psf.sickkids.on.ca

I recommend doing a time-course.  You should take aliquots a t=0 (no thrombin)
20 min, 40 min, 1 hr, 2, 4, 8 and you'll probably find out that all that
you can 
get cut will be cut within the 1st 2 hours, and might even find when the 
degradation of the peptide occurs.

m

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