The UGA codon in vitro: Chain termination and suppression

Abstract

UGA suppressor-dependent mutants of bacteriophage f2 were isolated at low frequency after fluouracil mutagenesis. One was mutant in the viral polymerase cistron, and six of the remainder were defective in the maturation protein cistron. No coat protein mutants have been found. RNA extracted from the polymerase mutant (op 9) was used to stimulate the in vitro synthesis of phage-specific products in extracts derived from su − E. coli. The labeled products were compared with those stimulated by wild-type f2 RNA, and with those stimulated by RNA from sus 10, an amber polymerase mutant of f2. RNA from op 9 stimulates the synthesis of 20 to 30% as much viral polymerase as does f2 RNA, whereas sus 10 RNA does not stimulate the formation of any polymerase. Each mutant RNA acts as a template for the synthesis of a peptide which is not found when f2 or the other mutant RNA is used. When su − extracts are supplemented with su I + soluble RNA, sus 10 stimulates the formation of intact polymerase. The addition of su UGA + soluble RNA to op 9 RNA-stimulated su − extracts results in an increased formation of intact polymerase and a decreased synthesis of the polymerase fragment. It is concluded that the UGA codon, like the UAG codon, causes premature termination of the polypeptide chain, and that termination can be suppressed by the appropriate sRNA.