Re: [ccp4bb] Are these xtal conditions worth optimizing?

2012-06-08 Thread mjvdwoerd 
Hi Christine,

I would try to optimize both conditions, provided that you do not have 
contradicting information (like a diffraction pattern that shows a small 
lattice - salt). Have you tried an additive screen? Have you tried adding a 
detergent? 

My experience is mixed with things of that nature - sometimes you can get 
better looking crystals, sometimes not. And you should always keep in mind 
that looks and content (and subsequently diffraction quality) are two 
different matters. Once I was taught a rule that in the case of protein-DNA 
complexes, the quality of diffraction is inversely proportional to the 
prettiness of the crystals and then I proved that rule wrong with 
nice-looking crystals that did diffract well. But it is true that crystals do 
not need to be pretty to be useful. So by all means, please do test diffraction.

Good luck.

Mark



-Original Message-
From: Harman, Christine christine.har...@fda.hhs.gov
To: CCP4BB CCP4BB@JISCMAIL.AC.UK
Sent: Wed, Jun 6, 2012 3:53 pm
Subject: [ccp4bb] Are these xtal conditions worth optimizing?


 
Hi All,
I have these very weird drops that I found from screening (please find pictures 
attached).  I am not sure if they are worth optimizing.  I am very interesting 
to know your opinions of what you think of thesedrops could be (are these what 
you call spherulites?).  If you think these conditions are worth optimizing, 
I welcome any ideas on how to optimize.  I have done some optimization and 
still get the same result which is many of these weird things growing 
throughoutthe drop and with no sharp edges, and sometimes a skin forms after 
2weeks (with the sodium malonate conditions only).   I have also opened the 
drop and poked around to see if it is phase separation and this things are 
definitely solid and slightly-very mushy. I haven't had a chance to check for 
diffraction, but will be very soon.   Both of these drops contain the same 
protein preparation of a Fab/peptide complex @ ~5mg/mL in buffer containing 
0.1M Sodium Acetate pH 5, 150mM NaCl. I appreciate any advice, thoughtsor 
comments that you could provide.
 
Peace,
Christine

Re: [ccp4bb] Are these xtal conditions worth optimizing?

2012-06-07 Thread Enrico Stura 

Dear CCP4bb,

1) Check the bottom of the tube from which the protein was taken. Round
objects could be sephadex beads.

2) Touch the object with a cat wisker. Does it break up easily?
Yes: probably protein crystals. If it is an oil instead of a solid object  
you have

your answer.

3) If these objects break up easily can you use them as seeds?
If so, it is well worth optimizing!!

Birefringence is also useful, but you need to know your salts. Some are  
difficult
to distinguish from small (less than 60 aa) proteins that can also be  
strongly birefringent.
Birefringence is useful in particular if you check how fast the color  
changes as you rotate the
cross-polarizer. Portein crystals tend not to change color as fast as salt  
crystals do. Sephadex

beads are not birefringent.

Enrico.


On Thu, 07 Jun 2012 00:10:47 +0200, Jacob Keller  
j-kell...@fsm.northwestern.edu wrote:



Birefringence?

JPK

On Wed, Jun 6, 2012 at 4:52 PM, Harman, Christine 
christine.har...@fda.hhs.gov wrote:



Hi All,
I have these very weird drops that I found from screening (please find
pictures attached).  I am not sure if they are worth optimizing.  I am  
very
interesting to know your opinions of what you think of these drops  
could be

(are these what you call spherulites?).  If you think these conditions
are worth optimizing, I welcome any ideas on how to optimize.  I have  
done

some optimization and still get the same result which is many of these
weird things growing throughout the drop and with no sharp edges, and
sometimes a skin forms after 2weeks (with the sodium malonate conditions
only).   I have also opened the drop and poked around to see if it is  
phase
separation and this things are definitely solid and slightly-very  
mushy.  I

haven't had a chance to check for diffraction, but will be very soon.
Both of these drops contain the same protein preparation of a  
Fab/peptide

complex @ ~5mg/mL in buffer containing 0.1M Sodium Acetate pH 5, 150mM
NaCl. I appreciate any advice, thoughts or comments that you could  
provide.


Peace,
Christine











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e-mail: est...@cea.fr Fax: 33 (0)1 69 08 90 71