In compactly folded RNAs, coordination or hydrogen bonding of cations in specific sites is a potentially important aspect of the tertiary structure. NH4+ specifically stabilizes the tertiary structure of a conserved, 58-nt fragment of the large subunit ribosomal RNA, as judged in two ways: a melting transition associated with tertiary interactions is sharpened and stabilized more effectively by NH4+ than by any alkali metal cation, and the affinity of the RNA fragment for ribosomal protein L11 or the antibiotic thiostrepton is approximately 10-fold stronger when measured in NH4+ than in Na+. The dependence of the melting temperature on NH4+ concentration shows that a single bound ion is responsible for these effects. The requirement of different ribosome functions for NH4+ suggests that other such sites exist in ribosomal RNAs.