Inter-sulfhydryl distances in plasma fibronectin determined by fluorescence energy transfer: effect of environmental factors.
Inter-sulfhydryl distances in plasma fibronectin determined by fluorescence energy transfer: effect of environmental factors.
复制标题
通过荧光能量转移测定血浆纤连蛋白中的巯基间距离：环境因素的影响。
DOI：
10.1021/bi00465a030
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发表时间：
1990
期刊：
影响因子：
2.9
通讯作者：
Lai,CS
中科院分区：
文献类型：
--
作者：
Wolff,CE;Lai,CS
材料和方法sfn在ge-latin-Sepharose 4B亲和柱上从威斯康星州东南部血液中心的新鲜冷冻人血浆中纯化（Engvall & Ruoslahti, 1977）。用巯基试剂标记前后，用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）常规检测蛋白质的完整性和纯度。荧光探针7V-(l-pyrenyl)雄性亚胺(P), 7-（二乙胺）-3-（4，-马来酰苯基）-4-甲基香豆素(C)和荧光素5-马来酰亚胺(F)购自Molecular probes （Eugene， OR）。带有表面羧基的聚苯乙烯乳胶珠购自Polysciences （Warrington， PA）。本研究中使用的珠子直径为0.5¿tm。三（羟甲基）氨基甲烷（Tris），苯基甲基磺酰氟（PMSF）， 1 -[3-（二甲基胺）丙基]-3-乙基碳二亚胺盐酸盐（碳二亚胺），胰蛋白酶（牛胰腺）和肝素（猪）
Materials and MethodsFn was purified from fresh-frozenhuman plasma, obtained from the Blood Center of Southeastern Wisconsin, on a ge-latin-Sepharose 4B affinity column (Engvall & Ruoslahti, 1977). The integrity and purity of the protein, before and after labeling with the sulfhydryl reagents, were routinely examined with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The fluorescence probes 7V-(l-pyrenyl) male-imide (P), 7-(diethylamino)-3-(4,-maleimidylphenyl)-4-methylcoumarin (C), and fluorescein 5-maleimide (F) were purchased from Molecular Probes (Eugene, OR). Polystyrene latex beads with surface carboxyl groups were purchased from Polysciences (Warrington, PA). The beads used in this study had a diameter of 0.5¿ tm. Tris (hydroxymethyl) aminomethane (Tris), phenylmethanesulfonyl fluoride (PMSF), l-[3-(di-methylamino) propyl]-3-ethylcarbodiimide hydrochloride (carbodiimide), trypsin (bovine pancreas), and heparin (porcine