lac Operator nucleosomes. 1. Repressor binds specifically to operator within the nucleosome core.

Area of Science:

• Molecular Biology
• Biochemistry
• Genetics

Background:

• Histones package DNA into nucleosomes, influencing gene regulation.
• The lac repressor controls the lac operon by binding to the lac operator DNA sequence.

Purpose of the Study:

• To investigate the simultaneous binding of lac repressor and histone octamers to DNA.
• To determine if histone binding affects lac repressor recognition of the lac operator.

Main Methods:

• Formation of artificial nucleosomes and core particles using DNA fragments and core histones.
• Characterization of these reconstitutes.
• Lac repressor binding assays, including operator-dependent binding and inhibition by IPTG.
• Sedimentation studies to assess histone octamer displacement.
• Cross-linking experiments using formaldehyde.

Main Results:

• Lac repressor specifically binds to the lac operator within artificial nucleosomes and core particles.
• Histone octamers are not displaced by repressor binding, allowing simultaneous association.
• Binding is operator-dependent and specifically inhibited by IPTG (isopropyl thiogalactoside).
• Cross-linking experiments confirm simultaneous binding.
• Inferred that the operator DNA's binding surface faces outward, away from the histone core.

Conclusions:

• The lac repressor and histone octamer can bind simultaneously to the same DNA region.
• Nucleosome structure allows for specific DNA-protein interactions without displacing core histones.
• The accessibility of the operator DNA sequence is crucial for simultaneous binding.