发信人: crystalmaker (风雨无阻), 信区: Biology
标 题: Re: protein crystallography
发信站: Unknown Space - 未名空间 (Sat Jun 19 23:43:57 2004) WWW-POST

Sorry for being so late. These days I just was too busy.

When I was in China we seldom us Screen kits becau it is too expensive,
even we had these kits. Some people made these kits themselves. Now in
US these kits are so easy to get.
I usually try Hampton and Wizard kits first. sometimes good luck, but often
nothing.
First check sample purity and stability. it is important.
For precipitation, check it carefully. What kinds of precipitant it is.
sometimes precipitant is better than clear drops.
Besides vary pH and concentrations, I try organic reagents a lot, and
some did give good results. Like Dioxane, isopropenol, DMF, etc.
I also try detergents for precipitation. I found it very useful in my
proteins when I was in china. I even made a small paper on using detergents
in crystallization (a very low journal).

Sometimes you may need to think about making different constucts if the sample
is expressed. This will be a lot of work to do.

and do not forgot ask experts' opinion at ccp4bb,cnsBB and O mailing list,
etc.
There are more experts there than here.

And sometimes, some proteins is too difficult to get crystals. I know some
proteins that have been found >10 years, but still no strucutures.

At last, crystallization is Luck+experience+knowledge. if it is too difficult,
then it is difficult. Like the monomer Insulin, people spent >30 years to
get it.

【 在 capry (IA) 的大作中提到: 】
: Thank you for reply.
:
: How about talking about some trick? I just started crystal screening for a
new
: bifunctional protein. Most of the results so far are precipitates. So how to
: move on by this frustrating reults?
:
: 【 在 nohate (无恨) 的大作中提到: 】
: :
: : me2
: :
: : 【 在 crystalmaker (风雨无阻) 的大作中提到: 】
: : : I am one. see my username.
: : :
: : :
: : : 【 在 capry (IA) 的大作中提到: 】
: : : : 版上有作蛋白质结晶方面的吗？也许我们可以交流一下。：）
: : : :
: : : :
: : :
: :
:
:

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