Some other things to try:
1. Co-crystallize with a ligand
2. crystallization with lipid cubic phase or bicelles
3. limited proteolysis to define a rigid core
ho
-
Hi all,
I got a crystal of one membrane protein (~60kD) from Na/K phosphate
condition (
Hi Qiangmin,
All the comments and references that were already mentioned to you are
excellent,
I would stress 3 points.
1- The detergent.
A clear distinction should be made between the detergent used for
extraction/solubilization and the detergent (or cocktail of
detergents/lipids) used for cryst
I cannot say what you have - running a gel, MS, etc of a washed crystal
could confirm what you have.
What you did not indicate is if your lack of diffraction was of frozen
or unfrozen crystals. I have seen too many cases where it is the cryo
condition killing diffraction. So if you have not tried
I cannot say what you have - running a gel, MS, etc of a washed crystal
could confirm what you have.
What you did not indicate is if your lack of diffraction was of frozen
or unfrozen crystals. I have seen too many cases where it is the cryo
condition killing diffraction. So if you have not tried
When diffraction fails - one can always look at the crystals with a UV
microscope (assuming you have Trp) - they should light up. Or if its
possible - harvest them, wash and run a gel. (or if its detergent a TLC)
Joe
2010/9/1 wu donghui
> It does not look like protein crystal and IZIT stainin
It does not look like protein crystal and IZIT staining is not reliable in
determining protein or other. Mostly it is like detergent or PEG crystal or
quasi-crystal.
Good luck
Wu
2010/8/31 qiangm zhang
> Hi all,
>
> I got a crystal of one membrane protein (~60kD) from Na/K phosphate
> co
I have had a lot of problems of my own with poorly diffracting (or not at all)
membrane protein crystals. After a previous discussion here, I summarised the
suggestions I got in this ccp4 user wiki;
http://strucbio.biologie.uni-konstanz.de/ccp4wiki/index.php/Improving_crystal_quality
There are
Yea, Parveen has actually asked about this here a year ago and I found this
discussion quite useful indeed:
www.mail-archive.com/ccp4bb@jiscmail.ac.uk/msg11717.html
and I think we all get tons of these crystals especially in conditions with
PEGs.
There are lots of things you can try and have been
Hi,
The picture looks like detergent crystals, specially DDM crystals. I have same
crystals in many conditions.
With regards,
Parveen
Hello Qiangmin,
Here are links to a few web resources that have tips for membrane protein
crystallization that may help with your optimization strategy:
http://kb.psi-structuralgenomics.org/update/2009/06/full/th_psisgkb.2009.25.html
http://web.emeraldbiosystems.com/blog/bid/33916/8-Practical-
On Tue, 2010-08-31 at 11:57 -0500, Jacob Keller wrote:
> I just don't want this guy to get misled into perhaps wasting
> months/years on something not particularly promising.
Trouble is, of course, that one never knows if a particular trick will
work this time. We routinely get PEG/fluoride salt
I have to say that in my limited experience with membrane protein
crystallization, these liquid crystal / spherulite type things are very
common, and seldom turn into anything useful. Perhaps others on the list
more experienced than I can corroborate or refute this. I just don't want
this guy to ge
Well said. I've seen three cases by now when switching to a homologue
from a different organism led to solving a structure (and way too many
cases when crystals just did not diffract, either at all or well
enough :).
On Tue, 2010-08-31 at 18:48 +0300, Tommi Kajander wrote:
> Or might be worth goi
Or might be worth going back to the drawing board to design more
constructs (and check them around the same conditions), thermostable
homologs etc.. what about reductive methylation, anyone had luck with
membrane proteins?
tommi
Quoting "Ed Pozharski" :
Unfortunately, some crystals don't
Unfortunately, some crystals don't diffract at all. You may want to try
to 100% verify that it's protein either by SDS-PAGE or mass-spec
(100x100x100 micron crystal could contain ~0.5mcg of protein, so you my
need to use silver staining). If it is, I'd consider trying to get
diffracting crystals
15 matches
Mail list logo