Re: [ccp4bb] How to remove nucleic acid contamination for crystallizing zinc finger protein

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CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999

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Subject: Re: How to remove nucleic acid contamination for crystallizing zinc finger protein
From: Ana Silva silva {- at -} YSBL {- dot -} YORK {- dot -} AC {- dot -} UK
Date: 2007-07-05

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Subject: Re: How to remove nucleic acid contamination for crystallizing zinc finger protein
From: William Scott wgscott {- at -} CHEMISTRY {- dot -} UCSC {- dot -} EDU
Date: 2007-07-05

Subject: Re: How to remove nucleic acid contamination for crystallizing zinc finger protein
From: Pavan p {- dot -} vaidyanathan {- at -} UMIAMI {- dot -} EDU
Date: 2007-07-05

You could also try polyethyleneimine (Polymin P) precipitation to
precipitate out your DNA. Add Polymin P dropwise to a final
concentration of 0.24% to your cell lysate while stirring in the cold
for an hour. This should precipitate out your DNA. Then centrifuge
your lysate and use the relevant fraction (supernatant or pellet) for
further purification.

Pavan

On 7/5/07, Ana Silva wrote:
> Hi,
>
> How much salt do you have in your protein buffer?
> I would try to increase the salt concetration, during purification.
>
> Hope it helps.
> Ana
>
>
>
> tony_htc@126.com wrote:
> >
> > Dear all,
> >
> > Sorry for the off-topic question.
> >
> > I am purifying a zinc finger transcription factor for crystallization.
> > The protein appeared as a single band on SDS-PAGE (MW 44KD) after NTA
> > chelating column, but its OD280/OD260 ratio is as high as 1.0. So I
> > doubt the protein is nucleic acid contaminated, probably because of
> > the zinc finger. I tried to remove the nucleic acid by Mono Q and
> > Superdex 75 pg, but failed. So could any one recommend some method to
> > remove the nucleic acid during protein crystallization, esp. zinc
> > finger protein? Any experience or references will be appreciated.
> >
> > Thanks a lot!
> >
> > Tiancen Hu
> >
> > Shanghai Institute of Materia Medica
> >
> >
> >
> > ------------------------------------------------------------------------
> > Ò»Æð À´£¬150 Íò ÈË Í¬ Ê± ÔÚ Íæ µÄ ÃÎ »Ã Î÷ ÓÎ
> >
>

--
Pavan
http://umsis.miami.edu/~pvaidyan

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CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999