DOMAIN-STRUCTURE OF THE RIBOZYME FROM EUBACTERIAL RIBONUCLEASE-P

Large RNAs can be composed of discrete domains that fold independently . One such ''folding domain'' has been identified previously in the ri bozyme from Bacillus subtilis ribonuclease P (denoted P RNA). This dom ain contains roughly one-third of all residues, Folding of an RNA cons truct consisting of the remaining two-thirds of B. subtilis P RNA was examined by Fe(II)-EDTA hydroxyl radical protection. This molecule fol ds into the proper higher-order structure under identical conditions a s the full-length P RNA, suggesting the presence of a second folding d omain in B. subtilis P RNA. Folding analysis of the Escherichia coli P RNA by hydroxyl radical protection shows that this P RNA is completel y folded at 5-6 mM Mg2+. I, order to analyze the structural organizati on of folding domains in E. coli P RNA, constructs were designed based on the domain structure of B. subtilis P RNA. Fe(II)-EDTA protection indicates that E. coli P RNA also contains two folding domains. Despit e the significant differences at the secondary structure level, both P RNAs appear to converge structurally at the folding domain level. The pre-tRNA substrate, localized in previous studies, may bind across th e folding domains with the acceptor stem/3'CCA contacting the domain i ncluding the active site and the T stem-loop contacting the other. Bec ause all eubacterial P RNAs share considerable homology in secondary s tructure to either B. subtilis or E. coli P RNA, these results suggest that this domain structure may be applicable for most, if not all, eu bacterial P RNAs. Identification of folding domains should be valuable in dissecting structure-function relationship of large RNAs.