Biopolymers and cell. 1985. Volume 1. № 4. 183 - 193

Yu. P. Semenkov, E. M. Makarov, S. V. Kirillov

QUANTITATIVE STUDY OF INTERACTION OF DEACYLATED tRNA WITH THE P, A AND E SITES OF ESCHERICHIA COLI RIBOSOMES

Summary

    The association constants of deacylated tRNAPhe for the P site of 70S ribosomes were measured in the presence and absence of poly(U), and at different Mg²⁺ concentrations and temperatures. The numbers of Mg²⁺ ions involved in this interaction were determi¬ned: 4.4+0.3 (+poly(U) and 2.7+0.3 (—poly(U). The association constant of tRNA^Phe for the A site was found to be 5·10⁵M^-1, much lower than that for the P site (2·10⁹M^-1), at 15 mM Mg²⁺, 200 mM NH₄⁺ and 25 °C. The independence of the E-site binding of tRNA^Phe on the messenger RNA was confirmed under these medium conditions. Based on the data obtained the affinity orders of aminoacyl-, peptidyl-tRNA and deacylated tRNA for the P, A and E sites are determined, and a possible mechanism of involvement of the E site in the translocation process is discussed.