INTER-RNA INTERACTION AND COMMUNICATION OF PHAGE ^o29 PRNA IN THE FORMATION OF HEXAMERIC COMPLEX AS A DNA TRANSLOCATION MOTOR

Peixuan Guo, Chaoping Chen, Kyle Garver, Mark Trottier and Chunlin Zhang
Department of Pathobiology and Purdue Biochemistry & Molecular Biology Program, Purdue University, West Lafayette, IN 47907

Linear ds-DNA viruses including bacteriophage ^o29 package their DNA into a preformed capsid during replication. A 120-base RNA (pRNA) encoded by ^o29 has a novel and essential role in genomic DNA translocation. The pRNA secondary structure has been partially solved and two functional domains have been identified. Homogeneous pRNA purified from a single band from denaturing gels showed six bands in native gels after incubation with magnesium, suggesting the presence of pRNA oligomers, including a potential hexamer. Atomic force microscopy also implied pRNA oligomer formation. Mutant pRNAs with intermolecular rather than intramolecular loop pairing were all inactive individually. Mixing of two, three and six inactive mutant pRNAs resulted in full DNA packaging activity as long as the interlocking hexameric ring can be predicted to form by base pairing of mutated loops. These results strongly suggested that six pRNAs formed a hexameric ring by inter-molecular interaction of two single-stranded RNA loops, and pRNAs interact and communicate with each other during DNA packaging through base pairing. This pRNA was absolutely required for procapsid- and DNA-dependent ATPase activity. Magnesium and ATP induced a conformational change in the pRNAs. These data, together with our previous published results showing that six pRNAs work sequentially during ^o29 DNA translocation, implying that ^o29 pRNA interacts intermolecularly to form a hexamer as part of a DNA translocation motor, and may have something in common with protein enzymes, such as DNA helicases. Formation of hexameric RNA also provides a model for the study of intermolecular interactions of RNA.