Transient-State Kinetic Analysis of the Ribosome Reaction Pathway

Dr. Marina V. Rodnina, Institute of Physical Biochemistry, University of Witten/Herdecke, Germany

Protein synthesis from amino acids in the cell is performed on ribosomes, large ribonucleoprotein particles that consist of several RNA molecules and over 50 proteins. The ribosome is a molecular machine that selects its substrates, aminoacyl-tRNAs, very rapidly and accurately and catalyses the synthesis of peptides from aminoacyl-tRNA. Recently, enormous progress has been made in understanding the ribosome by analysing its functions using rapid kinetics methods. Kinetic measurements showed that the ribosome not only bound correct substrates more tightly than the incorrect ones (stability discrimination), but also accelerated the rate of forward reactions for correct, rather than for incorrect tRNA (induced fit). This entirely new concept provided the basis for the interpretation of the interactions and conformational changes in the crystal structured of ribosomes and cryo-EM reconstructions of ribosome complexes. After the tRNA substrates are bound, the ribosome employs entropic catalysis to accelerate peptide bond formation by positioning substrates, reorganizing water in the active site, and providing an electrostatic network that stabilizes reaction intermediates. Proton transfer during the reaction appears to be promoted by a concerted shuttle mechanism that involves ribose hydroxyl groups on the tRNA substrate.

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