Detergent Desorption of Membrane Proteins Exhibits Two Kinetic Phases.
Detergent Desorption of Membrane Proteins Exhibits Two Kinetic Phases.
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DOI：
10.1021/acs.jpclett.8b00549
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发表时间：
2018-04-19
期刊：
影响因子：
--
通讯作者：
Movileanu L
中科院分区：
文献类型：
--
作者：
Wolfe AJ;Gugel JF;Chen M;Movileanu L
洗涤剂分子与水不溶性膜蛋白的逐渐解离最终导致蛋白质聚集。然而，该过程的时间依赖性轨迹仍然不明确，因为大多数光谱和量热技术的信噪比因溶液中蛋白质聚集体的存在而急剧下降。在这里，我们表明，通过使用稳态荧光偏振（FP）光谱，可以在洗涤剂浓度低于临界胶束浓度（CMC）的情况下实时检查蛋白质-洗涤剂复合物（PDC）的解离。本文提供了洗涤剂单体从膜蛋白解离的两个不同阶段共存的实验证据。我们首先注意到缓慢的去污剂预去溶剂化过程，伴随着 FP 各向异性相对适度的变化，表明蛋白质胶束中存在少量解离的去污剂单体。该预去溶剂化阶段之后是快速去污剂去溶剂化过程，其中 FP 各向异性的重大变化凸显了这一过程。这些阶段的持续时间和速率取决于去垢剂浓度和界面 PDC 相互作用。这种方法的进一步发展可能会导致创建一种新的半定量方法来评估蛋白质胶束的缔合和解离动力学。
Gradual dissociation of detergent molecules from water-insoluble membrane proteins culminates in protein aggregation. However, the time-dependent trajectory of this process remains ambiguous, because the signal-to-noise ratio of most spectroscopic and calorimetric techniques is drastically declined by the presence of protein aggregates in solution. Here, we show that by using steady-state fluorescence polarization (FP) spectroscopy, the dissociation of the protein-detergent complex (PDC) can be inspected in real time at detergent concentrations below the critical micelle concentration (CMC). This article provides experimental evidence for the coexistence of two distinct phases of the dissociations of detergent monomers from membrane proteins. We first noted a slow detergent predesolvation process, which was accompanied by a relatively modest change in the FP anisotropy, suggesting a small number of dissociated detergent monomers from the proteomicelles. This predesolvation phase was followed by a fast detergent desolvation process, which was highlighted by a major alteration in the FP anisotropy. The durations and rates of these phases were dependent on both the detergent concentration and interfacial PDC interactions. Further development of this approach might lead to the creation of a new semi-quantitative method for the assessment of the kinetics of association and dissociation of proteomicelles.
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