Abstract 2224 Bacterial and phage DNA structuring proteins control topoisomerase activity to promote replication

Discovery of the double-helical structure of the DNA led immediately to the recognition that the intertwined parental DNA strands must be untwisted for replication to occur. This DNA strand separation leads to formation of overtwisted or positively supercoiled DNA ahead of the replication fork. For replisomes to progress, positive supercoils must be removed by type II topoisomerases (DNA gyrase and topo IV in bacteria). The molecular mechanisms that regulate when, where, and the extent to which topoisomerases are active are poorly understood. We have described GapR, a protein that recognizes the shape of positive supercoiling and promotes replication by stimulating type II topoisomerase activity to remove these supercoils. Although GapR is found predominantly in alpha-proteobacteria, GapR homologs are found in many viruses (phages) that infect alpha-proteobacteria, suggesting that the phage GapR homologs function to promote phage infection. Here, we show using biochemical and genetic assays that phage GapR act analogously to its bacterial homolog, and recognizes positively supercoiled DNA and stimulates topoisomerase activity. We propose that phages have co-opted a bacterial replication promoting protein in order to target bacterial topoisomerase to phage DNA and promote phage replication. This remarkable cross-kingdom conservation of function raises the possibility that analogous regulators of positive supercoiling will be found in diverse species. This research was supported by a National Institutes of General Medical Science R00 grant (GM134153) to M.S.G.