PMID:8858581
| Citation |
Nègre, D, Bonod-Bidaud, C, Geourjon, C, Deléage, G, Cozzone, AJ and Cortay, JC (1996) Definition of a consensus DNA-binding site for the Escherichia coli pleiotropic regulatory protein, FruR. Mol. Microbiol. 21:257-66 |
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| Abstract |
The FruR regulator of Escherichia coli controls the initiation of transcription of several operons encoding a variety of proteins involved in carbon and energy metabolism. The sequence determinants of the FruR-binding site were analysed by using 6x His-tagged FruR and a series of double-stranded randomized oligonucleotides. FruR consensus binding sites were selected and characterized by several consecutive rounds of the polymerase chain reaction-assisted binding-site selection method (BSS) using nitrocellulose-immobilized DNA-binding protein. FruR was demonstrated to require, for binding, an 8 bp left half-site motif and a 3 bp conserved right half-site with the following sequence: 5'-GNNGAATC/GNT-3'. In this sequence, the left half-site AATC/ consensus tetranucleotide is a typical motif of the DNA-binding site of the regulators of the GalR-Lacl family. On the other hand, the high degree of degeneracy found in the right half-site of this palindrome-like structure indicated that FruR, which is a tetramer in solution, interacts asymmetrically with the two half-sites of its operator. However, potentially FruR-target sites showing a high degree of symmetry were detected in 13 genes/operons. Among these, we have focused our interest on the pfkA gene, encoding phosphofructo-kinase-1, which is negatively regulated by FruR. |
| Links | |
| Keywords |
Amino Acid Sequence; Bacterial Proteins/genetics; Bacterial Proteins/metabolism; Base Sequence; Binding Sites/genetics; Consensus Sequence; DNA Footprinting; DNA Methylation; DNA, Bacterial/chemistry; DNA, Bacterial/genetics; DNA, Bacterial/metabolism; Deoxyribonuclease I; Escherichia coli/genetics; Escherichia coli/metabolism; Escherichia coli Proteins; Genes, Bacterial; Molecular Sequence Data; Operon; Phosphofructokinase-1/genetics; Polymerase Chain Reaction; Promoter Regions, Genetic; Repressor Proteins/genetics; Repressor Proteins/metabolism; Sequence Homology, Amino Acid; Sequence Homology, Nucleic Acid |
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