CHARACTERIZATION OF SPERMIDINE BINDING TO SOLUBILIZED PLASMA-MEMBRANEPROTEINS FROM ZUCCHINI HYPOCOTYLS

In this work [C-14]spermidine binding to total proteins solubilized fr om plasma membrane purified from zucchini (Cucurbita pepo L.) hypocoty ls was investigated. Proteins were solubilized using octyl glucoside a s a detergent. Specific polyamine binding was thermolabile, reversible , pH dependent with an optimum at pH 8.0, and had a K-d value of 5 mu M, as determined by glass-fiber-filter assays. Sephadex G-25 M gel-fil tration assays confirmed the presence of a spermidine-protein(s) compl ex with a specific binding activity. By sodium dodecyl sulfate-polyacr ylamide gel electrophoresis and native polyacrylamide gel electrophore sis of collected fractions having the highest specific spermidine-bind ing activity,:several protein bands (113, 75, 66, and 44 kD) were iden tified. The specificity of spermidine binding was examined by gel-filt ration competition experiments performed using other polyamines and co mpounds structurally related to spermidine. Partial purification on Se phadex G-200 led to the identification of 66- and 44-kD protein bands, which may represent the putative spermidine-binding protein(s) on the plasmalemma.