DNA-STRUCTURE IN THE NUCLEOPROTEIN COMPLEX THAT ACTIVATES REPLICATIONOF PHAGE-029

Initiation of phage Phi 29 DNA replication is activated by the viral p rotein p6 which forms a nucleoprotein complex at the replication origi ns, located at the linear genome ends. The complex consists of a DNA r ight-handed superhelix wrapped around a multimeric protein core. We ha ve determined the superhelical path of the DNA in the complex, measuri ng the change in linking number induced by the protein, the surface-re lated helical repeat and the compaction of the DNA. One superhelical t urn has approximate to 63 bp (2.6 p6 dimers). Furthermore, we have det ermined that the DNA binding domain of protein p6 is located at the N- terminal region, predicted to form an amphipathic alpha-helix. We have obtained, by site-directed mutagenesis, protein p6 mutants in the pol ar side of the putative helix in which their DNA binding and replicati on activation properties were impaired or undetectable, in agreement w ith in vivo results.