Initiation Factor IF‐3 and the Binary Complex between Initiation Factor IF‐2 and Formylmethionyl‐tRNA Are Mutually Exclusive on the 30‐S Ribosomal Subunit

Abstract

The formation of 30‐S initiation complexes depends strongly on initiation factor IF‐3; at molar ratios of IF‐3 to 30‐S ribosomes up to one a stimulation is observed, whereas at ratios higher than one, initiation complex formation declines strongly. The target of the observed inhibition of fMet‐tRNA binding at high concentrations of IF‐3 is the 30‐S initiation complex itself. On the one hand addition of IF‐3 to preformed 30‐S initiation complexes leads to a release of bound fMet‐tRNA which is linear with the amount of factor added, whereas no effect on isolated 70‐S initiation complexes is seen. The release of fMet‐tRNA from preformed 30‐S initiation complexes is accompanied by a release of IF‐2 in a one‐to‐one molar ratio which is in agreement with our previous findings showing that binding of fMet‐tRNA takes place via a binary complex: IF‐2 · fMet‐tRNA (Eur. J. Biochem. 66, 181 ‐ 192 and 77, 69–75). On the other hand increasing amounts of both IF‐2 and fMet‐tRNA relieve the IF‐3‐induced inhibition of 30‐S initiation complex formation. From these findings it is concluded that IF‐3 and the IF‐2 · fMet‐tRNA complex are mutually exclusive on the 30‐S ribosome. This implies that under our experimental conditions MS2 RNA binding precedes fMet‐tRNA binding if one accepts that the presence of IF‐3 on the 30‐S subunit is obligatory for messenger binding. Copyright © 1978, Wiley Blackwell. All rights reserved