purification for GST pull down - (Dec/23/2005 )

Hi everybody

I would like to perform a GST pull down of my protein from cell extracts. Most of the protocols for GST pull down begin with binding of a given amount of your purified protein with your target extract, and then bind and pull down with glutathione-agarose beads.

My questions are: Is the reduced-glutathione elution protocol compatible with a second binding of the GSTfusion protein to glutathione-beads? Should I dialyze the eluted protein in any special condition? Which other elution protocols are suitable for this GST pull down purpouse? Should I keep my fusion protein bound to the resin and incubate directly with the extract instead of eluting it?

Thanks everybody (and have a happy Christmas!!)

Miguelon

Hi,

I think I didn´t explained myself very well...

Is it possible to bind again a GST-fusion protein to a glutathion resin, after eluting with reduced-glutathione?

Which other methods do you know of eluting a GST fusion protein from a glutathion-resin?

Thanks again!!