Re: [ccp4bb] Protein-DNA complex crystallization

2011-11-21 Thread Phoebe Rice 
What is Kd?  

Also, in reply to earlier posts: it is sadly common in crystallizing large 
protein-DNA complexes to go through a couple dozen different duplexes and 
several dismally-diffracting crystal forms before finding a good one.

  Phoebe

>From: CCP4 bulletin board  (on behalf of umar farook 
>)
>Subject: [ccp4bb] Protein-DNA complex crystallization  
>To: CCP4BB@JISCMAIL.AC.UK
>
>   Dear All,
>   I have been trying to crystallize protein DNA
>   complex, but all the time i end up with DNA
>   crystals. Even i changed the length of DNA many
>   times but still no complex, DNA only crystallizes!
>   Does anybody has idea, why do DNA crystallize by
>   itself ? My protein behaves very nicely, Dynamic
>   Light Scattering always shows nice values implies
>   homogenous but once i tried to ran acidic native
>   page but it shows little bit aggregated. The protein
>   is highly hydrophilic and soluble, and has only
>   three cysteines, is it there any possibility of
>   aggregation due to cysteine, when overexpressed in
>   E.coli ? and one more thing i mixed protein and DNA
>   together and ran agarose gel to see any gel shift,
>   indeed there is a binding, but when i take the same
>   thing to set drops, only DNA crystals. Kindly
>   suggest me, what could be done.
>   Thanks & Regards,
>   Umar Farook.S

Re: [ccp4bb] Protein-DNA complex crystallization

2011-11-21 Thread sxn214 


Sent from my Verizon Wireless Phone

- Reply message -
From: "umar farook" 
Date: Mon, Nov 21, 2011 10:59 am
Subject: [ccp4bb] Protein-DNA complex crystallization
To: 

Dear All,

I have been trying to crystallize protein DNA complex, but all the time i
end up with DNA crystals. Even i changed the length of DNA many times but
still no complex, DNA only crystallizes! Does anybody has idea, why do DNA
crystallize by itself ? My protein behaves very nicely, Dynamic Light
Scattering always shows nice values implies homogenous but once i tried to
ran acidic native page but it shows little bit aggregated. The protein is
highly hydrophilic and soluble, and has only three cysteines, is it there
any possibility of aggregation due to cysteine, when overexpressed in E.*
coli* ? and one more thing i mixed protein and DNA together and ran agarose
gel to see any gel shift, indeed there is a binding, but when i take the
same thing to set drops, only DNA crystals. Kindly suggest me, what could
be done.


Thanks & Regards,
Umar Farook.S

Re: [ccp4bb] Protein-DNA complex crystallization

2011-11-21 Thread Francis E Reyes 
Curious, how did you assess that your crystals only have DNA?

F

On Nov 21, 2011, at 8:59 AM, umar farook wrote:

> Dear All,
> 
> I have been trying to crystallize protein DNA complex, but all the time i end 
> up with DNA crystals. Even i changed the length of DNA many times but still 
> no complex, DNA only crystallizes! Does anybody has idea, why do DNA 
> crystallize by itself ? My protein behaves very nicely, Dynamic Light 
> Scattering always shows nice values implies homogenous but once i tried to 
> ran acidic native page but it shows little bit aggregated. The protein is 
> highly hydrophilic and soluble, and has only three cysteines, is it there any 
> possibility of aggregation due to cysteine, when overexpressed in E.coli ? 
> and one more thing i mixed protein and DNA together and ran agarose gel to 
> see any gel shift, indeed there is a binding, but when i take the same thing 
> to set drops, only DNA crystals. Kindly suggest me, what could be done.
> 
> 
> Thanks & Regards,
> Umar Farook.S



-
Francis E. Reyes M.Sc.
215 UCB
University of Colorado at Boulder

[ccp4bb] Protein-DNA complex crystallization

2011-11-21 Thread umar farook 
Dear All,

I have been trying to crystallize protein DNA complex, but all the time i
end up with DNA crystals. Even i changed the length of DNA many times but
still no complex, DNA only crystallizes! Does anybody has idea, why do DNA
crystallize by itself ? My protein behaves very nicely, Dynamic Light
Scattering always shows nice values implies homogenous but once i tried to
ran acidic native page but it shows little bit aggregated. The protein is
highly hydrophilic and soluble, and has only three cysteines, is it there
any possibility of aggregation due to cysteine, when overexpressed in E.*
coli* ? and one more thing i mixed protein and DNA together and ran agarose
gel to see any gel shift, indeed there is a binding, but when i take the
same thing to set drops, only DNA crystals. Kindly suggest me, what could
be done.


Thanks & Regards,
Umar Farook.S