Composition and functional characterization of yeast 66S ribosome assemblyintermediates

The pathway and complete collection of factors that orchestrate ribosome as sembly are not clear. To address these problems, we affinity purified yeast preribosomal particles containing the nucleolar protein Nop7p and develope d means to separate their components. Nop7p is associated primarily with 66 S preribosomes containing either 27SB or 25.5S plus 7S pre-rRNAs. Copurifyi ng proteins identified by mass spectrometry include ribosomal proteins, non ribosomal proteins previously implicated in 60S ribosome biogenesis, and pr oteins not known to be involved in ribosome production. Analysis of strains mutant for eight of these proteins not previously implicated in ribosome b iogenesis showed that they do participate in this pathway. These results de monstrate that proteomic approaches in concert with genetic tools provide p owerful means to purify and characterize ribosome assembly intermediates.