The effects of an affinity-purified polyclonal antibody to Artemia salina ribosomal protein L5 on protein synthesis in vitro were examined. The antibody interacted with 60 S subunits more strongly than with 80 S ribosomes, and inhibited reassociation of ribosomal subunits to some extent at 5 mM-Mg2" but not at 10 mm. Polyphenylalanine synthesis in vitro at 10 mM-Mg2" was significantly inhibited, especially when the antibody was first preincubated with 60 S subunits prior to the assay. The incorporation of amino acid directed by globin mRNA was inhibited only when the preincubation with 60 S subunits was carried out. On the other hand, no effect was detected on elongation factor 2-and 60 S subunit-dependent uncoupled GTPase activity. These results suggest that L5 is probably located at or near the subunit interface and may play an important role in protein synthesis.
INTRODUCTIONThe large ribosomal subunits of both prokaryotes and eukaryotes contain 5 S RNA, which is known to be a complex formed with one or more ribosomal proteins (for review, see ref. [1] [5][6][7] or other methods [8][9][10]. In rat liver, its protein moiety is identified as protein L5 [7]. One of us has shown that L5 crosslinked with poly(A)-containing mRNA in the polysome and to globin mRNA in the 80 S initiation complex following ultraviolet irradiation [11,12]. Furthermore, Uchiumi et al. [13] indicated that L5 crosslinked with small ribosomal subunit proteins S4 and S25 following treatment of 80 S ribosomes with 2-iminothiolane. These results suggest that rat liver L5 is probably located at the boundary between the large and small ribosomal subunits and interacts with mRNA.Recently, we identified the 5 S-RNA-binding protein of Artemia salina ribosomes, and showed immunological similarity between the proteins of Artemia and rat liver [14]. We therefore designated the protein as Artemia L5 (AL5) when compared with rat liver L5. In the present work, we used the affinity-purified antibody to AL5 and examined its effects on ribosomal functions in protein synthesis in vitro to elucidate the role of the 5 S-RNA-protein complex in ribosomes.