Kulcsár P, Prestwich G D
Department of Chemistry, State University of New York, Stony Brook 11794-3400.
FEBS Lett. 1988 Feb 8;228(1):49-52. doi: 10.1016/0014-5793(88)80582-9.
An [125I]iodinated juvenile hormone (JH) analog can be used as a sensitive and highly selective probe for the visualization of high-affinity, (JH)-specific binding proteins from insect hemolymph samples. The proteins can be detected in their native form using a two-dimensional (isoelectric focusing then native gradipore gel) separation of the crude protein mixture containing the 125I-labeled iodinated JH analog. The proteins can be transferred to activated glass fiber paper by electroblotting, and the location of the bound gamma-emitter can be found by exposure of the dried gel or the electroblot to X-ray film. The radiolabeled protein spot can be excised from the Coomassie-stained glass fiber paper and subjected directly to gas-phase N-terminal amino acid sequencing. This non-destructive, non-denaturing technique may have wide applicability in identifying and sequencing ligand-specific binding proteins in complex mixtures.
一种[125I]碘化保幼激素（JH）类似物可作为一种灵敏且高度选择性的探针，用于可视化昆虫血淋巴样品中的高亲和力、JH特异性结合蛋白。使用二维（等电聚焦然后是天然梯度孔凝胶）分离含有125I标记碘化JH类似物的粗蛋白混合物，可以以天然形式检测这些蛋白质。蛋白质可以通过电印迹转移到活化的玻璃纤维纸上，通过将干燥的凝胶或电印迹暴露于X射线胶片来确定结合的γ发射体的位置。可以从考马斯亮蓝染色的玻璃纤维纸上切下放射性标记的蛋白斑点，并直接进行气相N端氨基酸测序。这种非破坏性、非变性技术在鉴定和测序复杂混合物中配体特异性结合蛋白方面可能具有广泛的适用性。
