Puu G, Artursson E, Gustafson I, Lundström M, Jass J
Defence Research Establishment, Umeå, Sweden.
Biosens Bioelectron. 2000 Mar;15(1-2):31-41. doi: 10.1016/s0956-5663(00)00050-6.
Bacteriorhodopsin and the nicotinic acetylcholine receptor were biotinylated and reconstituted in lipidic membranes on silicon supports by fusion with proteoliposomes. The presence and distribution of the proteins were studied by binding with streptavidin. Radio-labelled streptavidin was employed for quantifying the amounts of protein remaining in the supported membranes after storage in buffer. The proteins within the membranes remained bound to the surface for weeks. The biological activity of reconstituted unlabelled receptor upon storage showed stability in membranes formed on silicon supports and a reduced stability when formed onto lipid monolayer covered supports. Atomic force microscopy studies on preparations in liquid showed bilayer structures but also attached, partly fused liposomes and membrane particles. In air, the surface was smoother and contained less of liposomes and more of stacked lipid layers. Preparations labelled with streptavidin conjugated to colloidal gold and imaged in air showed the proteins individually distributed, with no protein-rich patches or protein aggregates.
通过与蛋白脂质体融合，将细菌视紫红质和烟碱型乙酰胆碱受体进行生物素化，并重构于硅支持物上的脂质膜中。通过与链霉亲和素结合来研究蛋白质的存在和分布。使用放射性标记的链霉亲和素对在缓冲液中储存后支持膜中剩余的蛋白质量进行定量。膜内的蛋白质在表面保持结合状态达数周之久。重构的未标记受体在储存时的生物活性在硅支持物上形成的膜中表现出稳定性，而在脂质单层覆盖的支持物上形成的膜中稳定性降低。在液体中对制剂进行的原子力显微镜研究显示出双层结构，同时也有附着的、部分融合的脂质体和膜颗粒。在空气中，表面更光滑，脂质体较少，堆叠的脂质层较多。用与胶体金偶联的链霉亲和素标记并在空气中成像的制剂显示蛋白质呈单独分布，没有富含蛋白质的斑块或蛋白质聚集体。
