Specific DNA recognition by F Factor TraY involves beta-sheet residues.
The F Factor TraY protein is a sequence-specific DNA-binding protein required for efficient conjugal transfer. Genetic and biochemical studies indicate that TraY has two functional roles in conjugation. TraY binds to the PY promoter to up-regulate transcription of tra genes. TraY also binds to the plasmid origin of transfer (oriT), serving as an accessory protein in the nicking of F Factor in preparation for transfer. TraY is thought to belong to the ribbon-helix-helix family of transcription factors. These proteins contact DNA using residues of an antiparallel beta-sheet. We engineered and characterized six TraY mutants each having a single potential beta-sheet DNA contact residue replaced with Ala. Most TraY mutants had significantly reduced affinity for the TraY oriT binding site while possessing near wild-type stability and nonspecific DNA recognition. These results indicate that TraY beta-sheet residues participate in DNA recognition, and support inclusion of TraY in the ribbon-helix-helix family.
Department of Biology, The Johns Hopkins University, Baltimore, Maryland 21218, USA.
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D000595:Amino Acid Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D003001:Cloning, Molecular; D003227:Conjugation, Genetic / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D005144:F Factor / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011489:Protein Denaturation; D015202:Protein Engineering; D017510:Protein Folding; D017433:Protein Structure, Secondary
D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C063238:traY protein, E coli