The bicyclomycin sensitivities of 38 bicyclomycin-resistant mutants of transcription termination protein Rho and the location of their mutations support a structural model of Rho based on the F-1 ATPase

A total of 38 bicyclomycin-resistant mutants of Escherichia coli transcript ion termination protein Rho have been isolated. The locations of their muta tions identify the ATP-binding region as the functional domain inhibited by bicyclomycin. Strains containing the S266C, S266A and L208R Rho mutations are very resistant to bicyclomycin in vivo. In a similar way, the mutant Rh o proteins containing these mutations are very resistant to bicyclomycin in vitro. These data suggest that Ser266 and Leu208 might make direct contact with the antibiotic. These two residues are close to each other in the ter tiary structure of a model of Rho based on the alpha and beta subunits of t he F-1, ATPase, supporting the validity of the model. The strain containing the G337S Rho mutation also has high bicyclomycin resistance, and the prox imity of L208, S266 and G337 in the quaternary structure of the Rho model h as enabled a candidate bicyclomycin-binding pocket to be delineated. As a w hole, the bicyclomycin sensitivities of the mutants are consistent with the locations of their respective mutations in the model of Rho based on the F -1 ATPase, therefore supporting the emerging consensus model of Rho structu re. (C) 2000 Academic Press.