Marked stepwise differences within a common kinetic mechanism characterize TATA-binding protein interactions with two...

Area of Science:

• Molecular Biology
• Biochemistry
• Genetics

Background:

• Transcription initiation by RNA polymerase II requires TATA-binding protein (TBP) binding to promoter DNA.
• Understanding TBP-promoter interactions is crucial for deciphering gene regulation.

Purpose of the Study:

• To model and compare the interactions of Saccharomyces cerevisiae TBP with two strong promoters: E4 (TATATATA) and adenovirus major late (TATAAAAG).
• To elucidate the kinetic and thermodynamic mechanisms governing TBP binding and dissociation from these promoters.

Main Methods:

• Global analysis of kinetic and thermodynamic data.
• Fluorescence resonance energy transfer (FRET) measurements.
• Computational modeling of TBP-DNA interactions.

Main Results:

• A linear two-intermediate kinetic mechanism describes TBP binding to both promoters.
• Common features include tight binding, simultaneous DNA binding and bending, and stable intermediate conformers.
• Distinct dissociation pathways were observed: TBP-E4 dissociates via replacement and displacement, while TBP-adenovirus major late promoter dissociates via replacement only.
• Stepwise entropic and enthalpic compensations differ significantly between the two pathways.

Conclusions:

• TBP-promoter interactions share fundamental mechanistic steps but exhibit sequence-specific differences in dissociation.
• The study provides a foundation for comparing TBP interactions with consensus versus variant promoter sequences.