Hi Deliang,
My own experience with two membrane proteins suggests that His-tag
issues pertaining to soluble proteins also seem to apply to membrane
proteins.
In one case, for example, cleaving the N-terminal His-tag improved the
solubility of the protein dramatically. Interestingly, expression
levels of the protein without the His-tag were much lower than with it!
In the other case, we have a construct with a His-tag at the
N-terminus and another at the C-term. Both behave well when it comes
to solubility and stability but we thus far only have diffracting
crystals from the construct with the C-term His-tag. However, we have
not yet tried to remove the tags from those two constructs to see
whether that improves crystallizability.
My feeling is that if you do not have solubility/stability/aggregation
problems with your His-tagged membrane protein, then there is no
reason to expect lower success rates in crystallization trials. Screen
with it and hope for the best!
By the way, there is a recent paper on the effects of His-tags on
crystal structures of soluble proteins. It's worth a look.
Carson M et al. His-tag impact on structure.
Acta Crystallogr D Biol Crystallogr. 2007 Mar;63(Pt 3):295-301.
Best wishes
Savvas
----
Savvas N. Savvides
L-ProBE, Unit for Structural Biology
Ghent University
K.L. Ledeganckstraat 35
9000 Ghent, BELGIUM
Phone: +32-(0)9-264.51.24 ; +32-(0)472-92.85.19
http://www.eiwitbiochemie.ugent.be/units_en/structbio_en.html
Quoting deliang <[log in to unmask]>:
> Hi there,
>
> I purified a membrane protein with traditional His-tag on the C
> terminal. Before crystallization, I wonder how this tag may affect
> the result. Does anyone have experience that the removel of this
> tag may improve the result or not? or can provide some references
> which may give some statistic, like how many membrane proteins have
> been crystallized with or without His-tag?
>
> Thanks so much.
>
> Deliang
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