BIOPHYSICAL THEORY AND MODELING |
Distance-Restrained Docking of Rifampicin and Rifamycin SV
to RNA Polymerase Using Systematic FRET Measurements:
Developing Benchmarks of Model Quality and Reliability
Jennifer L. Knight 1, Vladimir Mekler 1, Jayanta Mukhopadhyay 1, Richard H. Ebright 1 and Ronald M. Levy 1*
1 Rutgers University
* To whom correspondence should be addressed. E-mail: ronlevy{at}lutece.rutgers.edu.
Submitted on August 2, 2004
Revised on September 16, 2004
Accepted on 2 November 2004
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Abstract |
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We are developing distance-restrained docking strategies for modeling macromolecular complexes that combine available high-resolution structures of the components and inter-component distance restraints derived from systematic fluorescence resonance energy transfer (FRET) measurements. In this paper, we consider the problem of docking small-molecule ligands within macromolecular complexes. Using simulated FRET data, we have generated a series of benchmarks that permit estimation of model accuracy based on the quantity and quality of FRET-derived distance restraints, including the number, random error, systematic error, distance distribution, and radial distribution of FRET-derived distance restraints. We find that expected model accuracy is 10 Å or better for models based on (i) >=20 restraints with up to 15% random error and no systematic error or (ii) >=20 restraints with up to 15% random error, up to 10% systematic error, and a symmetric radial distribution of restraints. Model accuracies can be improved to 5 Å or better by increasing the number of restraints to >=40 and/or by optimizing the distance distribution of restraints. Using experimental FRET data, we have defined the positions of the binding sites within bacterial RNA polymerase (RNAP) of the small-molecule inhibitors rifampicin (Rif) and rifamycin SV (Rif SV). The inferred binding sites for Rif and Rif SV were located with accuracies of, respectively, 7 and 10 Å relative to the crystallographically defined binding site for Rif (Campbell et al., 2001). These accuracies agree with expectations from the benchmark simulations and suffice to indicate that the binding sites for Rif and Rif SV are located within the RNAP active-center cleft, overlapping the binding site for the RNA-DNA hybrid.
Key Words:
Distance-restrained docking, FRET, Macromolecular assemblies, Modeling, RNA polymerase, Rifampicin
