Translation arrest by oligodeoxynucleotides complementary to mRNA coding sequences yields polypeptides of predetermined length

Abstract

We investigated the arrest of mRNA translation at predetermined sites by oligodeoxynucleotides complementary to defined coding sequences within a mRNA. An in vitro transcription and a wheat germ cell-free translation system were used for the synthesis of mRNA and protein, respectively. Oligodeoxynucleotides (10−, 15− and 20−mer) arrested polypeptide synthesis in a concentration-dependent manner at the site of their hybridization to the mRNA, as judged by the size of the translation products. A 5-mer oligodeoxynucleotide did not prevent synthesis of the full length protein. Ribosomes arrested by an oligodeoxynucleotide transiently stacked up and eventually disassembled. Upon dissociation of the ribosomes from the blocked site, nascent chains were released as peptidyl-tRNAs which in turn became rapidly converted to free polypeptide chains. None of these results was affected by the position within the reading frame to which the 3′ end of the oligodeoxynucleotide hybridized. The general applicability of translation arrest by oligodeoxynucleotides was demonstrated for different mRNAs. Only partial arrest of translation was obtained when oligodeoxynucleotides were used to arrest translation in the reticulocyte cell-free system.