Copyright X) 1993, American Society for Microbiology RNA Duplex Unwinding Activity of Poliovirus RNA- Dependent RNA Polymerase 3DP01

The ability of highly purified preparations of poliovirus RNA-dependent RNA polymerase, 3DPo1, to unwind RNA duplex structures was examined during a chain elongation reaction in vitro. Using an antisense RNA prehybridized to an RNA template, we show that poliovirus polymerase can elongate through a highly stable RNA duplex of over 1,000 bp. Radiolabeled antisense RNA was displaced from the template during the reaction, and product RNAs which were equal in length to the template strand were synthesized. Unwinding did not occur in the absence of chain elongation and did not require hydrolysis of the 'y-phosphate of ATP. The rate of elongation through the duplex region was comparable to the rate of elongation on the single-stranded region of the template. Parallel experiments conducted with avian myeloblastosis virus reverse transcriptase showed that this enzyme was not able to unwind the RNA duplex, suggesting that strand displacement by poliovirus 3DP01 is not a property shared by all polymerases. Poliovirus, the prototypic member of the Picomnaviridae family, has a single-stranded RNA genome of positive po-larity approximately 7,500 nucleotides (nt) long. The genome is covalently linked to a small viral protein, VPg, at its 5' terminus (9, 21) and includes a genetically encoded poly(A