Enzymatic Synthesis of Deoxyribonucleic Acid

Abstract

Abstract The Escherichia coli and T4 DNA polymerases do not extend chains in which the 3'-terminal nucleotide (primer terminus) is not paired with the template. By using synthetic double-stranded polynucleotides, the 3' → 5' exonuclease function of these polymerases was shown to be directed specifically against a mispaired or unpaired primer terminus. Chain extension of such termini begins only after all mispaired nucleotides have been removed and a basepaired terminus is reached. The latter is completely conserved while polymerization is maintained. These results suggest that the function of the 3' → 5' exonuclease activity of DNA polymerases is to remove mispaired nucleotides which have been incorrectly incorporated, thereby increasing the fidelity of template copying. The function of other E. coli exonucleases suggested by their specificities on polynucleotide substrates are trimming loose ends of DNA for exonuclease I and enlarging nicks and gaps within helical regions for exonuclease III.