Prot./DNA interaction and SDS-PAGE

Rogier rogier666 at
Fri Oct 13 04:27:31 EST 2000

Hi Emir,
SDS-PAGE will definitely destroy any protein-DNA interaction, unless the
binding is covalent.
6 histidines is about 0.6 kDa, you won't separate such a small size dif
on a gel. So you probably have two proteins. Unless you use a
high-percentage gel with lots of bisacrylamide, then you may pick up
different phosphorylation states of your protein.

To be really sure, cut out the two protein bands, cut with trypsin, run
on a high-percentage gel (or better, a tricine gel) and see if the
patterns are the same or not.

Good luck,

Rogier Stuger
Dept MicFizz, Free U of A
E rogier AT biogate DOT com (spammers will be killed)
do NOT use the address. I never read that one

> His6 DNA-binding protein shows 2 very close bands on SDS-PAGE.
> two forms, bound and non-bound to DNA? binding to DNA
> retained after boiling in SDS-PAGE buffer.expressed
> gene has second start codon after 6His
> Emir

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