On the fidelity of DNA replication.

Homogeneous DNA polymerase (“reverse transcriptase”) from avian myeloblastosis virus was assayed for exodeoxyribonuclease activity. The substrates were defined template. initiator complexes in which different radioactive nucleotides were present at the 3’-OH termini of the initiator. Even when the number of molecules of enzyme was equal to the number of initiator termini, there was no significant release of radioactivity with any of the template-initiator combinations tested. Under similar conditions, the nuclease activity associated with either Escherichia coli or T, DNA polymerases rendered more than 90% of the initiator termini acid-soluble. The ratio of exodeoxyribonuclease activity to protein with avian myeloblastosis DNA polymerase is less than 0.003% of that obtained with E. coli DNA polymerase I. Furthermore, avian myeloblastosis virus DNA polymerase failed to excise mispaired terminal nucleotides in both the presence and absence of polymerization. Avian myeloblastosis virus DNA polymerase utilizes mispaired initiator termini when complexed to ribohomopolymer and deoxyribohomopolymer templates as functional sites for chain propagation. The efficiency of utilization of the paired and mispaired termini is about equal in that the amount of synthesis and the size of the products formed are similar. The ability of the enzyme to utilize the mismatched termini as starting points for polymerization has been unambiguously demonstrated by transfer of 32P from the newly synthesized product to the mispaired 3’-terminal nucleotide on the initiator.