Studies on polynucleotides: LXXIII. Synthesis in vitro of polypeptides containing repeating tetrapeptide sequences dependent upon DNA-like polymers containing repeating tetranucleotide sequences: Direction of reading of messenger RNA

Abstract

The double-stranded DNA-like polymers, poly d-TATC:GATA ‡ and poly d-TTAC:GTAA, which contain in each strand repeating tetranucleotide sequences indicated by the nucleotide initials, have been used as templates for DNA-dependent RNA polymerase. By providing in the reaction mixture the three ribonucleoside triphosphates necessary for the transcription of only one of the two strands, the following four single-stranded ribopolynucleotides have been prepared: poly r-UAUC, poly r-GAUA, poly r-UUAC and poly r-GUAA. All of the ribopolynucleotides were shown to contain repeating tetranucleotide sequences by nearest-neighbor frequency analysis. In the cell-free amino acid-incorporating system from Escherichia coli B, poly r-UAUC and poly r-UUAC, each directed the incorporation of only four and three amino acids respectively. As expected from the three-letter, non-overlapping code, the polypeptidic product formed by poly r-UAUC was shown to have the repeating tetrapeptide sequence, Tyr-Leu-Ser-Ile, whereas the polypeptidic product from poly r-UUAC had the repeating tetrapeptide sequence, Leu-Leu-Thr-Tyr. With poly r-GUAA and r-GAUA as messengers, no acid-insoluble polypeptides were formed. The present results provide (1) further independent proof of the direction of reading of the messenger RNA and (2) confirmation of a total of ten codon assignments, including the nonsense triplets UAA and UAG, in the strain B of E. coli ‡ Abbreviations used: Poly d-TATC : GATA refers to the DNA-like polymer which contains the repeating units thymidylyl-dcoxyadenylyl-thymidylyl-deoxycytidylyl in one strand and deoxyguanylyl-deoxyaddenylyl-thymidylyl-deoxyadenylyl in the complementary strand. Poly d-TTAC : QTAA is a similar abbreviation. The ribopolymers are distinguished by the prefix r- immediately before the repeating unit and are single-stranded.