Structural basis of nucleotide-dependent conformational switching of a CBS-family protein on the archaeal 50S ribosome.

Ribosome, the universally conserved cellular machinery, orchestrates protein synthesis across all organisms. Ribosome-associated factors that regulate translation during biogenesis and stress adaptation remain underexplored in archaea due to limited structural data. Here we present eight cryogenic electron microscopy (cryo-EM) structures of the archaeal 50S ribosomal subunit at resolutions of 2.07-3.26 Å, revealing a previously uncharacterized cystathionine β-synthase (CBS) domain-containing protein positioned above the peptidyl transferase center, with an extended loop that inserts deeply into the peptide exit tunnel. Intriguingly, distinct adenosine derivatives occupy the ligand-binding pocket of the CBS protein in different structural states of the 50S subunit. Molecular dynamics simulations indicate that ATP binding induces a more contracted conformation of the extended loop, suggesting that nucleotide occupancy may modulate this protein's ribosome-association mode. Biochemical and bioinformatic analyses reveal that the extended loop is predominantly found in archaea, suggesting lineage-specific adaptations of ribosome-associated CBS proteins. Furthermore, the relatively sparse and dispersed distribution of rRNA modifications likely reflects organism-specific adaptations. Collectively, this work provides a structural framework for understanding how CBS-family proteins engage the archaeal ribosome and how their association may be tuned by cellular nucleotide status, offering insights into the molecular diversity of ribosome-associated factors and their roles in stress adaptation.