On 20 May 1997, David A. Elsemore wrote:
> Hello, I am looking for an explanation for an experimental result. I
> have recently looked for E. coli genes that when overexpressed provide
> resistance of E. coli to the herbicide glyphosate. Glyphosate targets
> the enolpyruvylshikimate-3-phosphate synthase. One clone that allowed E.
> coli to grow in the prescence of glyphosate contained the glucokinase
> locus.
We won't dwell on all the possible errors that could be leading to this
(e.g., Athel's "red herrings"). But there are some possible explanations
worth at least thinking about. Glyphosate is competitive with
phosphoenoyl pyruvate (PEP) in its inhibition of EPSP Synthase. We suspect
that a change in the PEP pool level is far more likely to effect
Glyphosate sensitivity than a general effect on flux to the aromatic amino
acid synthesis via promotion of glucose utilization. Remember that
shikimate is NOT competitive with Glyphosate on the enzyme, which is one
"reason" glyphosate works so well.
Consider some of the following possible things that can effect PEP
formation and/or utilization: The phosphotransferase system for sugar
import leads to glucose-6-phosphate directly with PEP as the original
phosphate donor. There are several proteins involved in this, and the
whole interesting system is one of the classic stories in microbial
biochemistry (early work was by Saul Roseman, see e.g. 1964 in PNAS 52,
1067-1074). Among other things, this transport system also seems to be the
primary transducer for "Catabolite Repression", in which a lack of free
glucose induces cAMP synthesis, which turns on expression of genes
involved in utilization of non-hexose carbon sources, etc. Consider also
possible deregulation of the first stages of glycolysis, without release
of pyruvate kinase from control, or direct inhibitory effects on pyruvate
kinase, leading to a higher steady state PEP level.
If you are at duPont's Experimental Station, you may want to talk to a
postdoc in Carl Falco's group in Ag Products, Sabine Epelbaum. She did her
PhD with us on interactions between the targets of herbicide inhibitors of
branched-chain amino acid synthesis and various other genes. Robert A.
(Bob) LaRossa, an excellent microbial genetecist/physiologist who
understands these things very well, is also at the Station. Give them
regards from us!
David Chipman and Ze'ev Barak
- - - - - - - - - - - - - - - - - -
David M. Chipman
Ze'ev Barak
Department of Life Sciences, Ben-Gurion University
POB 653, Beer-Sheva 84105 ISRAEL
Tel: {+972}-7-647 2646 Fax: {+972}-7-647 2890
Email: chipman at bgumail.bgu.ac.il
