Protection of avian myeloblastosis virus (AMV) DNA polymerase by substrates against heat inactivation.

Purified DNA polymerase from AMV was exposed to thermal inactivation in the presence and absence of various compounds required for maximal activity. A synergistic protective effect was noted when all the components were present. The enzyme showed a distinct preference for primed templates, especially for its native 70S RNA. Heat inactivation studies also suggested that the active sites for RNase H and that for the polymerase were not identical. It is well established that AMV DNA polymerase may initiate transcription of viral 70S RNA utilizing either a naturally occurring RNA or a synthetic oligodeoxynucleotide as a primer (1). As with most other DNA polymerases, the enzyme cannot synthesize DNA chains de novo using a single stranded template alone, since a 3'-OH containing primer molecule is required for initiation of synthesis. Thus addition of the primer oligo dT to AMV 70S RNA resulted in a six-fold stimulation in the template activity of the RNA (2), probably because of priming of the poly rA tract in the 70S RNA molecule (3). We have recently reported chemical modification of adenosine residues in poly rA (4) and AMV 70S RNA which resulted in their .inactivation as templates (2). The modified 70S RNA was also found to completely inhibit the transcription of unmodified 70S RNA. By contrast, inhibition was not observed when oligo dT primed 70S RNA was used as the template-primer, suggesting a preference by the enzyme for transcription of oligo dT primed regions of the 70S RNA (2). These initial observations prompted us to investigate further the effects of priming naturally occurring or synthetic templates on polymerase