Structure and conformational transitions of the of E. Coureplicative helicase DnaB protein hexamer - SsDNA complex

We present evidence that, in the presence of the ATP nonhydrolyzable analog AMP-PNP, the DnaB helicase binds polymer ssDNA with the site-size of 20± 3 nucleotides per protein hexamer. Studies of the 20-mer binding to the DnaB hexamer show that the hexamer has only a single, strong binding site for ssDNA. Photo-cross-linking experiments indicate that only a single subunit is primarily in contact with ssDNA. This surprisingly very low site-size of the large hexameric helicase-ssDNA complex, the existence of only a single, strong ssDNA binding site on the hexamer, and the results of photo-cross-linking experiments preclude the possibility of extensive wrapping of the ssDNA around the hexamer and formation of the complex in which all six protomers are simultaneously bound to ss nucleic acid. These results indicate that long-range allosteric interactions occur on the level of the quaternary structure of the hexameric enzyme, leading to the selection of a limited set of subunits as a binding site for ssDNA. We provide the first direct evidence of dramatic global conformational changes of the DnaB hexamer, induced by nucleotide cofactors and ssDNA binding, and the presence of multiple conformational states of the enzyme.