Hi Chiara,

according to my own experience, interaction between protein and DNA can be strong enough to "protect" DNA from DNAse action. 
To explain why with protease only the band disappeared, i need to know if the gel was run for long time, and if it's concentrate or not. Because if the DNA fragment is short, it passed trough the gel and you see nothing after.
The band that you see when you run without protease, maybe don't reflect the actual size of the putativ DNA fragment, but the "size" of the complex protein+DNA, in this case mostly the protein size.

Hope to help.

Nicolas


________________________________________
De : CCP4 bulletin board [[log in to unmask]] de la part de Chiara Rapisarda [[log in to unmask]]
Envoyé : lundi 29 juin 2015 10:48
À : [log in to unmask]
Objet : Re: [ccp4bb] Interesting DNA contamination

Hi,

I found that my protein bound ethidium bromide in an agarose gel. I tested that by treating my protein with protease and DNAse in two different tubes and running a gel. The band in the agarose gel disappeared only when the
protein was treated with protease. It is worth trying.
I hope that helps,

Chiara


Pramod,

You already got good suggestions on how to handle DNA contamination in protein preparations.

Let me point out briefly that you haven't demonstrated yet that your contamination is DNA.

I had the same observation when purifying UvsX. A very persistent and strong contamination in all my preps at ~500kb. To test weather it was DNA or RNA I boiled the protein 30 minutes and incubated it with DNAase and RNAse and result was the same. I concluded it was neither RNA nor DNA and continued as if nothing had happened.

This publication is reporting the same observation:

Formosa and Alberts (1986) "Purification and characterization of the T4 bacteriophage uvsX protein." J Biol Chem. 1986 May 5;261(13):6107-18.

If you ever find out what it is that runs like 500kb DNA on Agarose, please let me know.
S.

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Date:    Sat, 27 Jun 2015 04:39:45 +0100
From:    Stefan Gajewski <[log in to unmask]<mailto:[log in to unmask]>>
Subject: Re: Interesting DNA contamination

Correction,

I meant to say 0.5kb, not 500kb

sorry for that.
S.

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Date:    Sat, 27 Jun 2015 16:52:06 +0000
From:    "Rose, Peter" <[log in to unmask]<mailto:[log in to unmask]>>
Subject: Postdoctoral Fellows in Big Data/Structural Bioinformatics at University of California, San Diego

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Peter Rose, Ph.D.
Site Head, RCSB Protein Data Bank West (http://www.rcsb.org<http://www.rcsb.org/>)
Principal Investigator, Structural Bioinformatics Laboratory (http://bioinformatics.sdsc.edu<http://bioinformatics.sdsc.edu/>)
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------------------------------

Date:    Sat, 27 Jun 2015 17:00:08 +0000
From:    "Rose, Peter" <[log in to unmask]<mailto:[log in to unmask]>>
Subject: Web Developers at RCSB PDB at University of California, San Diego

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End of CCP4BB Digest - 26 Jun 2015 to 27 Jun 2015 (#2015-182)
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