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Macromolecular binding equilibria in the lac repressor system: studies using high-pressure fluorescence spectroscopy.
C A Royer1, A E Chakerian, K S Matthews
1Department of Physics, University of Illinois, Urbana 61801.
Biochemistry
|May 22, 1990
Summary
High hydrostatic pressure causes lac repressor tetramers to dissociate into dimers, with significant volume changes observed. Inducer binding influences this dissociation, especially at higher temperatures.
Area of Science:
- Biochemistry
- Biophysics
- Molecular Biology
Background:
- Lac repressor protein regulates gene expression in E. coli.
- Protein subunit interactions are crucial for protein function and stability.
- High hydrostatic pressure is a tool to study molecular assembly and dissociation.
Purpose of the Study:
- To investigate the effect of high hydrostatic pressure on lac repressor tetramer dissociation.
- To analyze the influence of inducer binding on lac repressor subunit association.
- To determine the volume changes associated with lac repressor tetramer dissociation and operator complex dissociation.
Main Methods:
- Fluorescence polarization using a long-lived fluorescent probe (dansyl).
- High hydrostatic pressure application to protein samples.
- Spectroscopic analysis of fluorescence polarization changes.
Main Results:
- High hydrostatic pressure induces dissociation of the lac repressor tetramer into dimers, with a volume change of -82 mL/mol.
- Dissociation constant increases with temperature, indicating reduced tetramer stability at higher temperatures.
- Operator binding destabilizes the lac repressor tetramer, and pressure promotes dissociation of repressor-operator complexes.
Conclusions:
- High hydrostatic pressure perturbs lac repressor quaternary structure, favoring dimer formation.
- Inducer binding shows a temperature-dependent effect on tetramer stabilization.
- Pressure-induced dissociation of repressor-operator complexes is more pronounced than tetramer dissociation.