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zerosignal

Flexible linker proteins on SDS-PAGE

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Hello everyone,

 

I've made a recombinant protein with a long (GGGGS)6 linker between two domains, and refolded it from inclusion bodies by chemical refolding in the presence of a redox couple.

 

Under non-reducing SDS-PAGE, I get a fairly broad (maybe 5-10kDa range) coomassie-staining band. This band is much more compact under reducing conditions (and shifted upwards, which I would expect).

 

My concern is whether the apparent divergence in mass of the band is indicative of incomplete oxidative refolding, or whether this is a property of having a long flexible linker between the domains.

 

Does anyone here have experience of expressing proteins with long flexible linkers and if so, did you observe a similar pattern by SDS-PAGE?

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