FUNCTIONAL OLIGOMERIZATION OF POLIOVIRUS RNA-DEPENDENT RNA-POLYMERASE

Using a hairpin primer/template RNA derived from sequences present at the 3' end of the poliovirus genome, we investigated the RNA-binding a nd elongation activities of highly purified poliovirus 3D polymerase. We found that surprisingly high polymerase concentrations were require d for efficient template utilization. Binding of template RNAs appeare d to be the primary determinant of efficient utilization because bindi ng and elongation activities correlated closely. Using a three-filter binding assay, polymerase binding to RNA was found to be highly cooper ative with respect to polymerase concentration. At pH 5.5, where bindi ng was most cooperative, a Hill coefficient of 5 was obtained, indicat ing that several polymerase molecules interact to retain the 110-nt RN A in a filter-bound complex. Chemical crosslinking with glutaraldehyde demonstrated physical polymerase-polymerase interactions, supporting the cooperative binding data. We propose a model in which poliovirus 3 D polymerase functions both as a catalytic polymerase and as a coopera tive single-stranded RNA-binding protein during RNA-dependent RNA synt hesis.