Modulation of theEscherichia coliTryptophan Repressor Protein by Engineered Peptides

Abstract

We have used theE. colitryptophan repressor (TrpR) as a model protein for modulation by engineered peptides bothin vivoandin vitro.The tryptophan operator promoter-lacZ reporter system was used to investigate thein vivoability of several synthetic peptides to modulate TrpR function. GMSA (gel mobility shift analysis) was used to study thein vitroability of the peptides to modulate binding of the TrpR protein to the operator DNA. Peptides WRW, DRW, DW, RW enhanced TrpR binding to the operatorin vivoat 100 μM concentrations. The same peptides enhanced TrpR binding to the operatorin vitroat 1 mM concentrations. The peptide RRW reduced TrpR binding to the tryptophan operator bothin vivoandin vitro.Thus the peptide RRW acted more as an inducer than corepressor. The peptide WR could neither enhance nor impede binding between TrpR and the operatorin vivoorin vitro,suggesting that the presence of a carboxyl tryptophan residue may be necessary for binding to the TrpR protein. Thin layer chromatography was used to ensure that the peptides had not been subject to proteolysis during thein vitrogel mobility shift assays.