Cooperative interactions in the system ribosomes-ribosomal protein S1-polynucleotide triplets.

D J Goss, L J Parkhurst, A M Mehta, A J Wahba

Biochemistry 1984 Dec 18

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Association equilibria have been determined in the ternary system uridyl triplets (T)-ribosomal protein S1 (S)-ribosomes (Rb) depleted of S1 at 6 and 10 mM Mg2+. For 1:1 stoichiometry of reactants, four thermodynamically independent equilibria characterize the ternary system. The binary interaction Rb + T was studied by following the fluorescence quenching of labeled ribosomes by added T. The Rb + T association constant for UpUpUp triplets was 10-20-fold greater than for ApUpG triplets. The interaction Rb + S was studied by following the changes in fluorescence anisotropy when labeled S1 reacted with ribosomes. The remaining two independent equilibrium constants (for S + T and RbT + S) were obtained from fits to observed anisotropy measurements when varying amounts of T were added to a solution of ribosomes and fluorescently labeled S1. This indirect procedure allows one to measure S + T binding, an association that is difficult to determine directly. Over the concentration interval 5-10 mM Mg2+, the association constant for Rb + S increases with the sixth power of [Mg2+], whereas the association constant for S + T decreases approximately 2-fold as Mg2+ is increased from 6 to 10 mM Mg2+. T binds to Rb more tightly at 10 mM than at 6 mM Mg2+. When S1 is bound to Rb, however, at 10 mM Mg2+ the binding constant for T is decreased 10-fold and the Mg2+ dependence is reversed. These interactions can be described in terms of coupling free energies. For the ternary complex, three linearly independent coupling free energies can be written.(ABSTRACT TRUNCATED AT 250 WORDS)

Citation

D J Goss, L J Parkhurst, A M Mehta, A J Wahba. Cooperative interactions in the system ribosomes-ribosomal protein S1-polynucleotide triplets. Biochemistry. 1984 Dec 18;23(26):6522-9