The DNA target of the trp repressor

Area of Science:

• Molecular Biology
• Structural Biology
• Genetics

Background:

• High-resolution X-ray crystallography of the trp repressor and trp operator complex revealed unexpected features.
• These features led to claims that the DNA fragment used was not the true operator, questioning the crystal structure's accuracy.
• An alternative operator sequence was proposed based on mutational studies and gel retardation analysis.

Purpose of the Study:

• To re-examine the sequence consensus in trp repressor-regulated (trpR) promoters.
• To analyze existing mutagenesis experiments to validate or refute the alternative operator model.
• To clarify the specific DNA sequence recognized and preferred by the trp repressor.

Main Methods:

• Re-analysis of sequence consensus in trpR-repressible promoters.
• Analysis of published mutagenesis experiments on trp repressor-DNA interactions.
• In vitro trp repressor-DNA binding assays using novel DNA constructs and full-length promoter sequences.

Main Results:

• The traditional operator sequence is fully consistent with mutagenesis data and stereochemical analysis.
• The proposed alternative operator sequence is stereochemically inconsistent with known interactions.
• In vitro binding assays demonstrate the trp repressor preferentially binds the traditional operator sequence.

Conclusions:

• The crystal structure of the trp repressor-operator complex accurately portrays a specific interaction.
• The traditional trp operator sequence is the authentic target recognized by the trp repressor.
• This finding resolves conflicting data and confirms the structural model's biological relevance.