EVIDENCE FOR A SINGLE NAPHTHYLPHTHALAMIC ACID-BINDING SITE ON THE ZUCCHINI PLASMA-MEMBRANE

The binding of [2,3,4,5,(n)-H-3]N-1-napthylphthalamic acid ([H-3]NPA) to zucchini (Cucurbita pepo L.) plasma membranes was examined in detai l using two different filtration assays and the results were rigorousl y analyzed by saturation curves, double-reciprocal plots, Scatchard pl ots, Hill plots, and the computer program Ligand (P.J. Munson, D. Rodb ard [1980] Anal Biochem 107:220-239). To facilitate these analyses, a new assay that allows rapid and quantitative analysis of [H-3]NPA bind ing with high reproducibility and ease of manipulation has been develo ped. These detailed kinetic analyses indicate that only one binding si te for [H-3]NPA (K(d) = 16 nm) was associated with the zucchini plasma membrane. Analysis of [H-3]NPA dissociation by several auxin transpor t inhibitors revealed similar dissociation constants with both plasma and microsomal membrane. Collectively, these data indicate the presenc e of only one binding site for NPA associated with the zucchini plasma membrane.