tod.lorenz at ghawk.com (Tod Lorenz) wrote:
> Could you explain what the Brent 2-hybrid system is for those
> that are not familiar with what it is and what it is used for?
> More details and particulars may help us solve the mystery....
The Brent two-hybrid system is generally called the Interaction
Trap. While the basic principles between the two systems are
similar, the actual components are completely different. Whereas
the Fields' two-hybrid uses the GAL4 DNA binding domain, the Brent
system uses LexA, a bacterial DNA protein with no known activity in
yeast. Whereas the two-hybrid uses the GAL4 activation domain or in
some cases the VP16 activation domain in the transactivator vector,
the Brent system uses B42, a bacterially derived "acid blob" that is
again completely heterologous to yeast. As a result of the elimination
of GAL4 components from DNA binding and activation domains, the
Brent system has the advantage that the library vector can be placed
under the control of the GAL1 inducible promoter, which removes a lot
of "Background" problems in library screening. In general, some
proteins of interest can be studied better using the Fields' system,
while others work better in the Brent system. I don't want this post to
run on forever: relevant references are
Golemis, E.A., Gyuris, J. and Brent, R. .(1994).
Two Hybrid Systems/Interaction Traps. in
Current Protocols in Molecular Biology,
F. M. Ausubel, R. Brent, R. Kingston et al, eds.
(New York, John Wiley and Sons), 13.14.1-13.14.17.
Gyuris, J., Golemis, E.A., Chertkov, H. and Brent, R. (1993)
Cdi1, a human G1 and S phase protein phosphatase that associates
with Cdk2. Cell 75: 791-803.
In further comment on Heidi Moss' post, the strain she was using is
not known to be prone to generation of ADE mutations, so the mystery
is still out there.....