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5a-formylbicyclomycin: studies on the bicyclomycin-Rho interaction
F Vincent1, W R Widger, M Openshaw
1Department of Chemistry, University of Houston, Texas 77204-5641, USA.
Biochemistry
|August 5, 2000
Summary
A new bicyclomycin derivative, 5a-formylbicyclomycin, irreversibly inactivates the rho transcription termination factor. This provides insights into bicyclomycin
Area of Science:
- Molecular Biology
- Biochemistry
- Microbiology
Background:
- Bicyclomycin is a commercial antibiotic targeting the rho transcription termination factor.
- Understanding the bicyclomycin-rho interaction is crucial for antibiotic development.
Purpose of the Study:
- To synthesize and characterize a novel irreversible inactivator of bicyclomycin, 5a-formylbicyclomycin.
- To elucidate the bicyclomycin-rho inactivation mechanism and identify the drug's binding site within rho.
Main Methods:
- Enzyme inhibition assays measuring rho poly C-dependent ATPase activity.
- Chemical modification studies involving imine formation and reduction to amine adducts.
- Mass spectrometry for stoichiometry determination and monomer exchange experiments.
Main Results:
- 5a-formylbicyclomycin is a more potent inhibitor (I50 = 35 microM) than bicyclomycin (I50 = 60 microM).
- Inhibition occurs via reversible, noncompetitive pathway with respect to ATP (Ki = 62 microM).
- Mass spectrometry indicates 5-6 bicyclomycin binding sites per rho hexamer; two modified subunits suffice for inactivation.
Conclusions:
- 5a-formylbicyclomycin acts as an effective irreversible inactivator of the rho transcription termination factor.
- The study defines the bicyclomycin binding site within the rho factor and its inactivation mechanism.