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Formation of 70S ribosomes: large activation energy is required for the adaptation of exclusively the small ribosomal subunit

Biophysical Chemistry, ISSN: 0301-4622, Vol: 96, Issue: 2, Page: 153-161
2002
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Article Description

Association of ribosomal subunits is an essential reaction during the initiation phase of protein synthesis. Optimal conditions for 70S formation in vitro were determined to 20 mM Mg 2+ and 30 mM K +. Under these conditions, the association reaction proceeds with first order kinetics, suggesting a conformational change to be the rate-limiting step. 70S formation separates into two sub-reactions, the adaptation of the ribosomal subunits to the association conditions and the association step itself. The activation energy of the process was determined to 78 kJ/mol and revealed to be required exclusively for the adaptation of the small subunit, rather than the large subunit or the association step. The presence of mRNA [poly(U)] together with cognate AcPhe-tRNA, accelerates the association rate significantly, forming a well-defined 70S peak in sucrose gradient profiles. mRNA alone provokes an equivalent acceleration, however, the resulting 70S couple impresses as an ill-defined, broad peak, probably indicating the readiness of the ribosome for tRNA binding, upon which the ribosome flips into a defined state.

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