Mass spectrometry reveals modularity and a complete subunit interaction map of the eukaryotic translation factor elF3.

The eukaryotic initiation factor 3 (elF3) plays an important role in translation initiation, acting as a docking site for several elFs that assemble on the 405 ribosomal subunit. Here, we use mass spectrometry to probe the subunit interactions within the human elF3 complex. Our results show that the 13-subunit complex can be maintained intact in the gas phase, enabling us to establish unambiguously its stoichiometry and its overall subunit architecture via tandem mass spectrometry and solution disruption experiments. Dissociation takes place as a function of ionic strength to form three stable modules elF3(c:d:e:l:k), elF3(f:h:m), and elF3(a:b:i:g). These modules are linked by interactions between subunits elF3b:c and elF3c:h. We confirmed our interaction map with the homologous yeast elF3 complex that contains the five core subunits found in the human elF3 and supplemented our data with results from immunoprecipitation. These results, together with the 27 subcomplexes identified with increasing ionic strength, enable us to define a comprehensive interaction map for this 800-kDa species. Our interaction map allows comparison of free elF3 with that bound to the hepatitis C virus internal ribosome entry site (HCV-IRES) RNA. We also compare our elF3 interaction map with related complexes, containing evolutionarily conserved protein domains, and reveal the location of subunits containing RNA recognition motifs proximal to the decoding center of the 405 subunit of the ribosome. [ABSTRACT FROM AUTHOR]