Tryptic core protein of lactose repressor binds operator DNA.

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• The lactose repressor protein controls the expression of genes involved in lactose metabolism in bacteria.
• Understanding repressor-DNA interactions is crucial for deciphering gene regulation mechanisms.

Purpose of the Study:

• To investigate the DNA-binding properties of the core lactose repressor protein, which lacks the N-terminal DNA-binding domain.
• To determine the contribution of the core protein to the overall binding energy and inducer-mediated repression.

Main Methods:

• Purification of the core lactose repressor protein using phosphocellulose chromatography.
• DNA binding assays to measure the dissociation constant of core protein binding to operator DNA.
• Competition studies with non-specific DNA to characterize the binding species.
• Kinetic experiments to assess the dissociation rate of core protein from operator DNA.

Main Results:

• The purified core protein binds to operator DNA with an apparent dissociation constant of 10(-7) M.
• Inducer presence decreases the binding of the core protein to operator DNA.
• Competition studies indicate that the binding species is not intact repressor or mixed tetramers.
• The core region contributes 40-50% of the binding energy to operator DNA.
• Inducer binding to core protein accounts for at least 60% of the free energy change observed for native repressor binding.

Conclusions:

• The core protein of the lactose repressor retains significant operator DNA binding capability.
• The N-terminal region is not solely responsible for repressor-operator interactions.
• These findings necessitate a re-evaluation of existing models for lactose repressor-DNA binding and gene regulation.