have lab, need enzyme

Jon Monroe monroejd at jmu.edu
Wed Oct 8 20:56:54 EST 1997


Thanks for all of the anatomy tips!  Please don't let this physiology
question interrupt...

I'm working with a student to develop a new enzyme lab for a Plant Phys
course.  What I would like to do is have students make crude extracts from
various plant sources, separate several isozymes relatively rapidly, then
do assays to compare the enzymes with respect to pH optimum, Km, etc., over
a 3 week period.  I've found that small concanavalin A-sepharose columns
will separate glycosylated from nonglycosylated proteins really quickly.  I
like the p-nitrophenyl-X based assays for their speed and simplicity.  Now
all I need is an enzyme for which +/- glycosylated forms exist.  We have
tried acid phosphatase, but, although it is potent, nearly all of it from
Arabidopsis shoots is glycosylated.  Ion exchange columns might be better
for separating different isozymes but would probably be too time consuming.
Does anyone know of an enzyme that fits (most of) these criteria?  One in
which different forms exist in the apoplast and the chloroplast would be
ideal...leading to simple fractionation experiments.

Thanks.

Jon

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