Inter-protein dependences during assembly of the 50S subunit from Escherichia coli ribosomes

Recently, we published an assembly map of the 50S ribosomal subunit from E. coli, incorporating 23S and 5S RNA and 26 out of the 32 L-proteins (Röhl and Nierhaus (1982) Proc. Natl. Acad. Sci. U.S.A., in press). In this paper we present the experimental details which led to the compilation of this 50S assembly map. The following experimental strategy was pursued: In order to elucidate binding of a protein to 23S RNA or a protein-protein interdependence, 23S rRNA and a set of purified ribosomal proteins were incubated under reconstitution conditions, the RNA-protein complex was separated from the non-bound proteins by sucrose gradient centrifugation and the proteins contained in the complex were determined by means of gel electrophoresis.