Hexameric Rings of Escherichia coli RuvB Protein.

Abstract

The Escherichia coli RuvA and RuvB proteins mediate ATP-dependent branch migration of Holliday junctions during homologous genetic recombination. RuvA is a DNA-binding protein with high affinity for Holliday junctions, to which it directs RuvB (a DNA-dependent ATPase). Electron microscopic studies have shown that RuvB forms double hexameric rings on duplex DNA. To determine whether the rings are biologically active, the conditions required for their formation and activity have been analysed. The quarternary structure of RuvB appears to be dependent upon the binding of ATP, magnesium ions, and the presence of RuvA. In the presence of Mg²⁺ and ATP, RuvB forms hexamers; however, in the presence of Mg²⁺ alone, dodecamers were observed. Both forms of the protein are stable and have been isolated by gel filtration. Preformed dodecamers and, to a lesser extent, hexamers assembled in the absence of DNA lack ATPase activity. Maximal ATPase activity was observed when RuvB assembled directly on DNA in the presence of Mg²⁺ and ATP. Moreover, under these conditions, a direct interaction between RuvB hexamers and tetramers of RuvA was observed.