DENSITY GRADIENT STUDY OF VICTORIN-BINDING PROTEINS IN OAT (AVENA-SATIVA) CELLS

Victorin-binding proteins (VBPs) in oat (Avena sativa) cells were iden tified using native victorin and anti-victorin polyclonal antibodies. Homogenates of oat tissues were fractionated in continuous or disconti nuous sucrose density gradients or with an aqueous two-phase method, a nd covalent binding sites of victorin were detected by western blottin g. In a 20 to 45% (w/w) sucrose continuous density gradient, the 100-k D VBP was located in fractions of 37 to 44% sucrose, with a peak at 39 % sucrose. Based on marker enzyme assays, plasma membranes peaked at 3 9 to 41% sucrose, mitochondria peaked at 41%, but Golgi and endoplasmi c reticulum were in lower density fractions, peaking at 28 to 29% and 22 to 24% sucrose, respectively. The 100-kD VBP was not found in plasm a membranes purified by the aqueous two-phase method or in mitochondri a purified by discontinuous density gradient centrifugation. Victorin binding to 65- and 45-kD proteins was detected in all fractions in the continuous sucrose density gradients. The 65- and 45-kD proteins were both detected in purified plasma membranes, but only the 65-kD protei n was detected in purified mitochondria. The subcellular location of V BPs was the same in sensitive and resistant oat cells.