Binding of Rifampicin to Escherichia coli RNA Polymerase: Thermodynamic and Kinetic Studies

Abstract

INTRODUCTION
The antibiotic rifampicin is a specific inhibitor of bacterial RNA polymerases (Hartmann et al. 1967) which, unlike many other compounds that interfere with transcription (Kersten and Kersten 1974), acts by direct interaction with the enzyme.

The existence of a stoichiometric and remarkably stable complex of rifampicin and RNA polymerase has been established in many ways (Wehrli et al. 1968; Wehrli and Staehelin 1970, 1971; Neuhoff, Schill and Sternbach 1970; Eilen and Krakow 1973; Scheit and Stütz 1975; Stender, Stütz and Scheit 1975). An actively transcribing ternary complex consisting of enzyme, DNA and RNA, however, does not bind the drug (Eilen and Krakow 1973) and is consequently resistant to its inhibitory action (Sippel and Hartmann 1968). The precise step in the sequence of reactions involved in transcription (Goldthwait, Anthony and Wu 1970; Chamberlin 1974b) beyond which sensitivity is no longer manifested remains to be established. Thus in recent years it has been postulated that rifampicin prevents either (a) the binding and stabilization effect of the initiating purine triphosphate (di Mauro et al. 1969); (b) the formation of an active state of the binary complex required for initiation (Sippel and Hartmann 1970); (c) the formation of the first phosphodiester bond (So and Downey 1970); or (d) the formation of the second phosphodiester bond (Johnston and McClure, this volume).

Although bound rifampicin does not prevent the specific recognition of promoter sequences directed by the σ subunit (Hinkle, Mangel and Chamberlin 1972; Bordier 1974), physical studies have demonstrated interference with the binding of...