Mesorhizobia are a type of nitrogen-fixing plant-symbiont bacteria. The genes that encode N-fixation and symbiosis are present on a mobile genetic element called an Integrative and Conjugative Element (ICE). The ICE from Mesorhizobium japonicum R7A; ICEMlSymR7A, is the prototypical member of the family of “symbiosis ICEs” that forms a symbiotic relationship with legumes. The RdfS protein works with a recombinase to stimulate excision of the ICE from the bacterial chromosome. We have demonstrated, using Surface Plasmon Resonance and Electrophoretic Mobility Shift Assays, that RdfS binds a region of DNA called an attachment site (att) in vitro. We have additional evidence in vitro and in vivo to suggest that RdfS is working to regulate conjugation of the ICE, in conjunction with its excisive role. X-ray crystallographic structural resolution to 2.5 Å shows RdfS contains a winged helix-turn-helix DNA-binding domain with a highly disordered C-terminal tail, which we suspect allows it to regulate additional conjugative machinery in the ICEMlSymR7A system and is a defining feature which differs from its homologs. Unravelling the multi-faceted role of RdfS gives greater insight into the tightly regulated molecular machinery involved in ICE excision and conjugation; and may potentially uncover a novel-type of protein regulation in transposable elements and symbiosis ICEs.