Kinetic control of histidine-tagged protein surface density on supported lipid bilayers
Kinetic control of histidine-tagged protein surface density on supported lipid bilayers
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DOI：
10.1021/la703788h
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发表时间：
2008-04-15
期刊：
影响因子：
3.9
通讯作者：
Groves, Jay T.
中科院分区：
文献类型：
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作者：
Nye, Jeffrey A.;Groves, Jay T.
镍螯合脂质是锚定多组氨酸标记的蛋白质支持的脂质双层（SLB）的一般工具，但存在争议的蛋白质-脂质附着的稳定性。在这里，我们表明，螯合剂脂质是合适的锚，用于建立稳定的，生物活性的表面，但一个简单的朗缪尔模型是不足以描述他们的行为。从螯合剂脂质的解吸动力学受表面结合的价数控制：单核苷酸结合的蛋白质在几分钟内解吸（t（12）约为6分钟），而多核苷酸结合的物质保持结合数小时（t（12）约为12小时）。表面状态之间的演化是缓慢的，因此在实验时间尺度上不太可能达到平衡。然而，通过调节孵育条件，可以动力学控制每个物种的群体，从而在具有单一浓度的螯合剂脂质的SL 13上提供宽范围的蛋白质密度。我们提出了与特定的蛋白质密度功能化的SLB表面的组装指南，并证明了它们在形成混合免疫突触中的实用性。
Nickel-chelating lipids are general tools for anchoring polyhistidine-tagged proteins to supported lipid bilayers (SLBs), but controversy exists over the stability of the protein-lipid attachment. Here, we show that chelator lipids are suitable anchors for building stable, biologically active surfaces but that a simple Langmuirian model is insufficient to describe their behavior. Desorption kinetics from chelator lipids are governed by the valency of surface binding: monovalently bound proteins desorb within minutes (t(1/2)approximate to 6 min), whereas polyvalently bound species remain bound for hours (t(1/2)approximate to 12 h). Evolution between surface states is slow, so equilibrium is unlikely to be reached on experimental timescales. However, by tuning incubation conditions, the populations of each species can be kinetically controlled, providing a wide range of protein densities on SLl3s with a single concentration of chelator lipid. We propose guidelines for the assembly of SLB surfaces functionalized with specific protein densities and demonstrate their utility in the formation of hybrid immunological synapses.