PURIFICATION OF A PROTEIN FROM AGROBACTERIUM-TUMEFACIENS STRAIN A348 THAT BINDS PHENOLIC-COMPOUNDS

In order to induce tumours on dicotyledonous plants, the bacterium Agr obacterium tumefaciens needs to be able to sense signal molecules, i.e . phenolic compounds. In order to identify putative chemoreceptors or environmental sensors involved in vir gene induction, we undertook the purification of a phenol-binding protein by affinity chromatography o n a syringamide-Ultrogel A4 column equilibrated at pH 5.6. A mild extr action of bacterial proteins with a Tris/HCl buffer at pH 9.0 led to t he purification of a 39 kDa protein (Pbp39) with a pi of 4.3 after spe cific elution of the affinity matrix with sodium syringate. When the a ffinity chromatography was performed at neutral pH, barely any protein was isolated, indicating the importance of an acidic pH for optimal a ffinity. A microplate binding experiment revealed that both syringyl-b iotinylated-BSA and sinapyl-biotinylated-BSA bound at pH 5.6 to the pl ate coated with Pbp39.