F. Dye et Fm. Delmotte, PURIFICATION OF A PROTEIN FROM AGROBACTERIUM-TUMEFACIENS STRAIN A348 THAT BINDS PHENOLIC-COMPOUNDS, Biochemical journal, 321, 1997, pp. 319-324
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.