title: The two main states of the elongating ribosome and the role of the alpha-sarcin stem-loop structure of 23S RNA.
abstract: According to the allosteric three-site model of the elongation cycle the ribosome oscillates between two main-functional states, viz the pre-translocational state with occupied A and P sites (E site with low affinity) and the post-translocational state with occupied P and E sites (A site with low affinity). This proposition could be confirmed by a determination of the thermodynamic parameters. High activation-energy barriers were found between both states, namely about 90 kJ mol-1 at 15 mM Mg2+ for either transition (post----pre transition = A-site binding and pre----post transition = translocation). The various A-site states (binding of ternary complex, EF-Tu dependent GTP cleavage, peptide-bond formation) are not separated by significant activation-energy barriers. The rate-limiting step of the elongation cycle is A-site binding, and not translocation as assumed previously. The principal role of both elongation factors is the reduction of the respective activation-energy barrier, thus accelerating the rate of the elongation cycle by several orders of magnitude. Cleavage of a single phosphodiester bond after G2661 of 23S rRNA by the RNase alpha-sarcin abolishes the functions of both elongation factors on the ribosome. This observation implies that the alpha-sarcin stem-loop structure plays an important role in the ribosomal conformational changes involved in the allosteric transitions. Indeed we could demonstrate that suitable oligodeoxynucleotide probes complementary to the alpha-sarcin region induce a conformational change in the 50S subunits; this conformational change causes an irreversible dissociation of tightly coupled ribosomes upon sucrose-gradient centrifugation.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Max-Planck-Institut für Molekulare Genetik, Berlin, Germany.
authors.forename: K H
authors.initials: KH
authors.lastname: Nierhaus
authors.forename: S
authors.initials: S
authors.lastname: Schilling-Bartetzko
authors.forename: T
authors.initials: T
authors.lastname: Twardowski
mesh_terms: D000495:Allosteric Site; D001483:Base Sequence; D004722:Endoribonucleases; D005656:Fungal Proteins; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010441:Peptide Chain Elongation, Translational / Q000502:physiology*; D012338:RNA, Ribosomal, 23S / Q000737:chemistry*; D012260:Ribonucleases; D012270:Ribosomes / Q000737:chemistry*
chemical_list: D005656:Fungal Proteins; D012338:RNA, Ribosomal, 23S; C014801:alpha-sarcin; D004722:Endoribonucleases; D012260:Ribonucleases