The Role of the Codon and the Initiation Factor IF‐2 in the Selection of N‐Blocked Aminoacyl‐tRNA for Initiation

Abstract

Poly(uridylic acid) [poly(U)] and poly(xanthidylic acid) [poly(X)] strongly stimulate the IF‐2‐dependent binding of fMet‐tRNA to 30‐S ribosomal subunits from Escherichia coli [Van der Laken et al. (1979) FEBS Lett. 100, 230–234]. The N‐formylmethionine moiety is incorporated into poly(phenylalanine) upon subsequent addition of other components required for protein synthesis when poly(U) is used as template. This paper shows that N‐acetylated Phe‐tRNAPhe (AcPhe‐tRNA), but not Phe‐tRNAPhe or tRNAPhe, competes with fMet‐tRNA for binding to poly(U)‐programmed 30‐S ribosomal subunits. The two species of N‐blocked aminoacyl‐tRNA are bound to poly(U)‐programmed and poly(X)‐programmed 30‐S subunits in a ratio that is linearly dependent on the ratio of the two species added. With poly(U) as template there is no apparent preference for either fMet‐tRNA or AcPhe‐tRNA, whereas with poly(X) there is a 2–3‐fold preference for fMet‐tRNA. The initiation factor IF‐2, which is strictly required for the binding of N‐blocked aminoacyl‐tRNAs, has a higher affinity for Met‐tRNA than for AcPhe‐tRNA. It is concluded that (a) interaction of the 30‐S ribosomal subunit with poly(U) or poly(X) leads to IF‐2‐dependent binding of N‐blocked aminoacyl‐tRNA; (b) the initiation factor IF‐2 discriminates in favour of fMet‐tRNA; (c) the presence of the cognate codon discriminates in favour of the corresponding N‐blocked aminoacyl‐tRNA. Copyright © 1980, Wiley Blackwell. All rights reserved