Protein-RNA interactions in the RNase P holoenzyme from Escherichia coli》.pdf

J. Mol. Biol. (1988) 202, 835848 Protein-RNA Interactions in the RNase P Holoenzyme from Escherichia coli Agustin Vioque’t, John Amez’ and Sidney Altmanl ‘Department of Biology and 2Department of Molecular Biophysics and Biochemistry Yale University New Haven, CT 06520, U.S.A. (Received 2 November 1987, and in revised form 8 March 1988) The genes for the protein (C5 protein) and RNA (Ml RNA) subunits of Escherichia coli RNase P have been subcloned and their products prepared in milligram quantities by rapid procedures. The interactions between the two subunits of the enzyme have been studied in vitro by a filter-binding technique. The stoichiometry of the subunits in the holoenzyme is 1 : 1. The dissociation constant for the specific interactions of the subunits in the holoenzyme complex is -4 x 10-l’ M. C5 protein also interacts with various RNA molecules in a non-specific manner with a dissociation constant of 2 x 10-s to 6 x 10-s M. Regions of Ml RNA required for interaction with C5 protein have been defined by deletion analysis and footprinting techniques. These interactions are localized primarily between nucleoti