title: Kinetic and thermodynamic parameters for tRNA binding to the ribosome and for the translocation reaction.
abstract: Kinetic analyses of tRNA binding to the ribosome and of the translocation reaction showed the following results. 1) The activation energy for the P site binding of AcPhe-tRNA to poly(U)-programmed ribosomes is relatively high (Ea = 72 kJ mol-1; 15 mM Mg2+). If only the P site is occupied with deacylated tRNA(Phe), then the E site can be filled more easily with tRNA(Phe) (no activation energy measurable) than the A site with AcPhe-tRNA (Ea = 47 kJ mol-1; 15 mM Mg2+). 2) A ribosome with blocked P and E sites represents a standard state of the elongation cycle, in contrast to a ribosome with only a filled P site. The two states differ in that AcPhe-tRNA binding to the A site of a ribosome with prefilled P and E sites requires much higher activation energy (87 versus 47 kJ mol-1). The latter reaction simulates the allosteric transition from the post- to the pretranslocational state, whereby the tRNA(Phe) is released from the E site upon occupation of the A site (Rheinberger, H.-J., and Nierhaus, K. H. (1986) J. Biol. Chem. 261, 9133-9139). The reversed transition from the pre- to the posttranslocational state (translocation reaction) requires about the same activation energy (90 kJ mol-1). 3) Both elongation factors EF-Tu and EF-G drastically reduce the respective activation energies. 4) The rate of the A site occupation is slower than the rate of translocation in the presence of the respective elongation factors. The data suggest that the A site occupation rather than, as generally assumed, the translocation reaction is the rate-limiting step of the elongation cycle.
authors.affiliation: Max-Planck-Institut für Molekulare Genetik, Abteilung Wittmann, Berlin-Dahlem, Germany.
authors.forename: S
authors.initials: S
authors.lastname: Schilling-Bartetzko
authors.forename: A
authors.initials: A
authors.lastname: Bartetzko
authors.forename: K H
authors.initials: KH
authors.lastname: Nierhaus
mesh_terms: D001665:Binding Sites; D007700:Kinetics; D012360:RNA, Transfer, Phe / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*; D013696:Temperature; D013816:Thermodynamics
chemical_list: D012360:RNA, Transfer, Phe