Assembly map of the 50-S subunit from Escherichia coli ribosomes, covering the proteins present in the first reconstitution intermediate particle

Highly purified proteins and 23-S RNA from the 50-S subunit of Escherichia coli ribosomes were used to study the assembly dependences of the early assembly proteins. The proteins under observation and the RNA were incubated at 4 mM Mg2+ and 44 degrees C, the unbound proteins were separated by sucrose gradient centrifugation, the RNA . protein complex was precipitated with trichloroacetic acid, and the complex-bound proteins was identified by means of sodium dodecylsulfate gel electrophoresis. A systematic analysis led to the establishment of an assembly map including 17 proteins which represent the protein moiety of the first reconstitution intermediate particle.