Need meth. to distinguish endo/exo glucanase

[Seanna Annis]

Christian Archambault (popa0215 at PO-Box.McGill.CA) wrote:
: I would need a method to determine if the protein we purified is an endo or exo beta-1,3 glucanase.  Any suggestion ???

: Christian Archambault
: popa0215 at po-box.mcgill.ca

You can easily do this by viscosity.  There are a lot of papers on using 
the technique for determining endo and exo polygalacturonases and this 
should work on glucanases depending upon what type of glucanase you 
have.  If carboxy methyl cellulose is a substrate for your enzyme you can 
make a viscous solution add your enzyme and then measure the viscosity at 
set intervals after you added your enzyme then plot determine 
the change in viscosity for each time and plot this against the reducing 
sugar released by your enzyme at each time (ie an identical time course 
except determine the reducing sugars released)  If the released reducing 
sugars are very high with a 50 % change in the viscosity than you have an 
exo enzyme otherwise if there are low levels of reducing sugar you have a 
endo enzyme.   If you need more info I can provide it   otherwise" Hoffman 
and Turner, 1982 physiological plant pathology" is a paper describing the 
viscosity method among other methods for polygalacturonase activity 
determination.  You can use the same methods on a variety of degradative 
enzymes.  Good luck Seanna Annis.  sannis at uoguelph.ca