tac vs T7 promoter, which is stronger?
Dr. Said Dermime
Dear All,
Both tac and T7 are considered 'strong' promoters which are frequently
used for expression of heterologous proteins in E. coli. Has anybody
done quantitative work on the relative strength of these two using a
reporter gene? If yes then could you point me towards the paper, please.
thanks
Armin Sepp
as...@rpms.ac.uk
Dr. Peter Gegenheimer
> Dear All,
> Both tac and T7 are considered 'strong' promoters which are frequently
> used for expression of heterologous proteins in E. coli. Has anybody
> done quantitative work on the relative strength of these two using a
> reporter gene? If yes then could you point me towards the paper,
> please.
Generally the T7 promoter will appear to be "stronger" in that
you'll get much more protein made from a T7 overexpression system
that from a tac-driven system. With the Studier T7 system (pET
vector and chromosomally-located T7 RNAP) one can easily [i.e.,
there are many reports in the lit., and we get this level with most
of the proteins we've tried] express the T7-driven gene to >= 50% of
total cell protein. I've never seen such levels from normal tac/trc
promoters, but I don't know of a side-by-side comparison.
Two caveats: first, a system with dual tac promoters gave excellent
expression of glycogen phosphorylase (paper from Bob Fletterick's
lab a year or so back), and because the rate of RNAP initiation can
be fine-tuned by the concentration of IPTG, they were able to get
low induction over a long period (1-2 days), so the protein remained
soluble. All of our T7-expressed proteins, with the notable
exception of groE chaperonin(!), were made as inclusion bodies.
Second, the "strength" of a promoter ought properly to mean either
the binding affinity for RNAP, or the clearance rate (rate at which
bound RNAP "clears" the promoter, i.e., is converted from
"initiating" to "elongating" polymerase). The former can be compared
only with the same RNAP; the latter could be compared among any
promoter/polymerase combinations. But no, I don't have references at
hand...
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