Hrazdina G, Lifson E, Weeden N F
Arch Biochem Biophys. 1986 Jun;247(2):414-9. doi: 10.1016/0003-9861(86)90600-4.
Chalcone synthase was isolated from illuminated buckwheat (Fagopyrum esculentum M.) hypocotyls and purified to electrophoretic homogeneity by sodium dodecyl sulfate-polyacrylamide gel electrophoresis using (NH)4SO4 fractionation, gel filtration on AcA 44, ion exchange chromatography on DEAE-Bio-Gel, and HPLC on hydroxylapatite. The properties of the enzyme were pH optimum, 8.0; Mr approximately 83,000 +/- 1000; Mr subunit, approximately 41,500 +/- 500; isoelectric point, pH 5.2; Km, 1 X 10(-6)M for malonyl-CoA, and 0.6 X 10(-6) M for p-coumaryl-CoA. Buckwheat chalcone synthase used p-coumaryl-CoA as substrate and also utilized caffeyl-CoA and ferulyl-CoA at 20 and 80%, respectively, of the rate of p-coumaryl-CoA in the chalcone synthase reaction. Antibodies against the buckwheat chalcone synthase were developed in a New Zealand white rabbit and characterized for specificity by enzyme-linked immunosorbent assay, Ouchterlony double immunodiffusion, and Western blotting.
从光照下的荞麦（甜荞）下胚轴中分离出查尔酮合酶，并通过硫酸铵分级沉淀、在AcA 44上进行凝胶过滤、在DEAE-生物凝胶上进行离子交换色谱以及在羟基磷灰石上进行高效液相色谱，将其纯化至电泳纯。该酶的性质为：最适pH 8.0；相对分子质量约83,000±1000；亚基相对分子质量约41,500±500；等电点pH 5.2；对丙二酰辅酶A的米氏常数为1×10⁻⁶M，对对香豆酰辅酶A的米氏常数为0.6×10⁻⁶M。荞麦查尔酮合酶以对香豆酰辅酶A为底物，在查尔酮合酶反应中，对咖啡酰辅酶A和阿魏酰辅酶A的利用率分别为对香豆酰辅酶A反应速率的20%和80%。用新西兰白兔制备了抗荞麦查尔酮合酶的抗体，并通过酶联免疫吸附测定、双向免疫扩散和蛋白质印迹法对其特异性进行了鉴定。
