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Team:INSA-Lyon/Project/Stage3/Strategy/Theorycurli
From 2010.igem.org
Theory concerning Curli and OmpR
You can find below some explanations about the curli promoter and the ompR protein :
Curli promoter
The Curli promoter comes from the chassis E.Coli PHL1273. It is a kanamycin resistant chassis containing a plasmid with the full-length intergenic region between csgD and csgB genes (approximately 450 bp) including csgD and csgBA promoters. A GFP reporter gene is placed downstream the csgBA promoter (curli promoter), in order to quantify its activity. The promoter csgD is under the control of the protein ompR. This chassis is adherent but all the genes involved in adherence structure synthesis (csgA,B,E,F and G genes) are in the chromosomal DNA, not in the plasmid DNA.
The curli production involves two divergent operons:
In addition, temperature and speed shaking conditions are also important in curli production. The precise mechanism of these regulations is still unknown, but some publications mention the Crl protein to be involved in the temperature regulation.
We made some measures of fluorescence to verify the influence of the temperature, shaking speed and osmotic pressure. You can see the results here.
In conclusion, the csgD promoter is regulated by osmolarity, temperature and speed shaking. Consequently, the strong csgB promoter, called “curli promoter” is regulated by these three conditions. Each condition is necessary but not sufficient.
OmpR protein
The OmpR protein, in special osmolarity conditions detected by the captor EnvZ, is phosphoryled. This phosphorylation facilitates its DNA binding on the csgD promoter to activate the CsgD protein synthesis. But, in wild-type classical laboratory strains of E.coli K12, curli, a particular class of envelope organelles, are not synthesized at a level sufficient to allow adherence.
When these strains are maintained in continuous culture for long-term experiences or industrial processes, adherent mutant cells are generated. A regulatory mutation is necessary to activate curli expression and gain adherence. This point mutation affects the regulatory properties of the OmpR protein and causes overproduction of curli. This mutation results in the replacement of a leucine by an arginine residue at position 43 due to the replacement of a T base by an G one at position 234.
It seems that the point mutation on OmpR234 protein facilitate phosphorylation and consequently its DNA binding. Therefore, the CsgD protein synthesis is increased and consequently it increases the expression of the csgB and csgA genes and so the production of curli.
