best fusion tag for protein purification
peter.cherepanov at uz.kuleuven.ac.be
Fri Sep 6 03:05:38 EST 2002
Of course, there is no such thing as "the best tag" for purification. It
depends on your experience only and vectors that you have in the lab. His
tag is exceptional, because it allows purification in both native and
denaturing conditions. BTW, inclusion bodies are not always that bad:
sometimes it is even desirable to have them. When your protein falls out as
inclusion bodies you will have much higher expression levels; unstable
proteins can be fused to a less soluble protein (like LacZ) just for that.
Sometimes you can increase soluble protein expression inducing your culture
at lower temperatures (28C or even 20C).
It seems that chance to get soluble protein is higher with other tags then
His6. MBP, for example, is considered to be the best for solubilization
(published somewhere). However, I heard, that after removal of the MBP part,
your protein might as well precipitate...
I do not know, if it is possible to increase solubility of the protein by
changing the host bacteria; may be it is, if you switch to a lower
expressor, but it will be the same at lower temperatures with your current
and to add finally: the best tag- no tag!
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