Influence of his-tag on protein folding
"jphilo*NO SPAM12*" at earthlink.net
Thu Apr 1 00:00:50 EST 1999
When I was at Amgen we saw cases where His tags significantly altered
the conformation and thermal stability of proteins. I also know of
proteins where the tag dramatically reduces the biological activity (but
these are proteins where the amino terminal region is directly involved
in their activity).
A number of labs that use analytical ultracentrifugation have found that
the tagged proteins are much more prone to self-association and
aggregation. Sometimes this is mediated through trace metal ions in
solution which bridge between the His tags, but in other cases the
enhanced association is not reduced in the presence of metal chelators.
So yes, the tag can really change the properties of your protein.
Unfortunately even when you use the protease to clip off the tag you
still don't end up with a natural amino terminus, although for many
proteins this is 'good enough'.
'Hope this helps,
Alliance Protein Laboratories
*** remove NOSPAM12 from address before replying ***
Balraj Doray wrote:
> Hi all,
> I'm presently using the his-tag sequence from Novogen's pET19b which
> has 12 histidines(10 in tandem), 4 aspartates(enterokinase cleavage site), 2
> glycines, 2 serines, a methionine, a lysine and an isoleucine residue in the
> sequence upstream of my own protein but I'm not using the pET19b vector but
> cloned the fusion sequence into another vector which uses the lac p/o control.
> What I'm observing is this tag is making a vast difference to the
> folding/assembly characteristics of some of my mutant fusions, namely, the
> presence of the tag greatly facilitates assembly of my mutant proteins. I'm
> observing this phenomena in both E. coli and T.ni insect cells. Has anyone
> any information or ideas on how this tag may be influencing the favourable
> folding of my mutant proteins or is there anything in the literature along
> these lines. Any comment would be greatly appreciated. Thanks!
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