We have proposed that the ancient ribosome increased in size during early evolution by addition of small folding-competent RNAs. In this Accretion Model, small RNAs and peptides were subsumed onto subunit surfaces, gradually encasing and freezing previously acquired components. The model predicts that appropriate rRNA fragments have inherited local autonomy of folding and local autonomy of assembly with ribosomal proteins (rProteins), and that the rProtein and rRNA are co-chaperones. To test these predictions, we investigate the rRNA interactions of rProtein uL23 and its tail, uL23^tail, which is a β-hairpin that penetrates deep into the core of the large ribosomal subunit. In the assembled ribosome, uL23^tail associates with Domain III of the rRNA and a subdomain called "DIII^core". Here using band shift assays, fluorescence Job plots, and yeast three-hybrid assays, we investigate the interactions of rProtein uL23 and its tail with Domain III and with DIII^core rRNA. We observe rRNA₁-uL23^tail₁ complexes in the absence of Mg²⁺ ions and rRNA₁-uL23^tail_n (n > 1) complexes in the presence of Mg²⁺ ions. By contrast, the intact uL23 rProtein binds in slightly anticooperative complexes of various stoichiometries. The globular and tail regions of rProtein uL23 are distinctive in their folding behaviors and the ion dependences of their association with rRNA. For the globular region of the rProtein, folding is independent of rRNA, and rRNA association is predominantly by nonelectrostatic mechanisms. For the tail region of the protein, folding requires rRNA, and association is predominantly by electrostatic mechanisms. We believe these protein capabilities could have roots in ancient evolution and could be mechanistically important in co-chaperoning the assembly of the ribosome.

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