Dear Ruby,
With this information I can give some more concrete hints:
- Add the sugar (monosaccharide, disaccharide?) that the lectin is
supposed to bind to your crystallization trials.
- Try different protein concentrations: 10 mg/ml, 100 mg/ml. In the
latter case you may have to dilute your screens.
- Try the same lectin from different plants.
- Do you purify it from the plant? I once purified a plant protein and
had to be very careful for phenol hydroxylases and had to add something
like dithionite before putting the plant in the blender. Adding protease
inhibitors may not hurt as well. You may want to check the integrity of
your protein for proteolytic cuts (giving ragged N- or C-termini) and
for other modifications making your protein heterogeneous. Is it
glycosylated? In that case you may want to try to remove it with some
glycanase.
Best,
Herman
________________________________
From: CCP4 bulletin board [mailto:[email protected]] On
Behalf Of ruby singh
Sent: Saturday, May 05, 2012 8:54 AM
To: [email protected]
Subject: [ccp4bb]
On Sat, May 5, 2012 at 12:17 PM, ruby singh
<[email protected]> wrote:
Sir,
i m sorry for not giving full details of protein.
its plant lectin protein on which im working currently.
its sequence information is very little. crystallization has become near
to impossible. the protein is a tetramer( of 28KDa). its purification
is very simple...but for crystallization i tried many things nt working
though(Hampton Reserach-pegion and index/CSS1/JCSG etc..). need some
help. the protein conecentration used was 30mg/ml stock . most of the
crystallization condition shows precipitation. the set up was put at
20C. the protein is very stable in MilliQ water upto 100mg/ml
concentration with no precipitation.
the protein doesnt aggregate..doesnt loose activity even after
boiling at 80C.
getting its crystal is important for my thesis. so need help.
