Linkage between operator binding and dimer to octamer self-assembly of bacteriophage lambda cI repressor

Area of Science:

• Molecular Biology
• Biophysics
• Genetics

Background:

• Bacteriophage lambda cI repressor dimer binding to operators OR and OL regulates phage development.
• Cooperativity was attributed to repressor dimer self-assembly into tetramers and octamers.
• Previous work identified octamer as the predominant species in repressor self-association.

Purpose of the Study:

• To investigate the coupling between bacteriophage lambda cI repressor self-association and operator binding.
• To identify unfavorable contributions to cooperativity beyond simple repressor self-assembly.
• To elucidate the mechanisms moderating cooperative protein-DNA interactions.

Main Methods:

• Sedimentation equilibrium analysis to compare dimer-octamer association of repressor-OR1 complex versus free repressor dimer.
• Fluorescence anisotropy to study OR1 binding to free dimers and higher-order assembled dimers.
• Utilized single operator site oligonucleotides to isolate protein-mediated effects.

Main Results:

• A significant, salt-dependent unfavorable contribution arises from the coupling between repressor self-association and operator binding.
• This protein-mediated coupling significantly reduces cooperativity.
• An additional salt-independent unfavorable contribution, likely DNA-mediated, also moderates cooperativity.

Conclusions:

• Protein-mediated coupling between repressor self-association and DNA binding acts as a key moderator of cooperativity.
• Structural transitions in both protein and DNA contribute to moderating cooperative interactions.
• This mechanism may represent a general principle in cooperative protein-DNA interactions.