Internal Structure and Preferential Protein Binding of Colloidal Aggregates.
Internal Structure and Preferential Protein Binding of Colloidal Aggregates.
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DOI：
10.1021/acschembio.6b00791
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发表时间：
2017-01-20
影响因子：
4
通讯作者：
Shoichet BK
中科院分区：
文献类型：
--
作者：
Duan D;Torosyan H;Elnatan D;McLaughlin CK;Logie J;Shoichet MS;Agard DA;Shoichet BK
小分子的胶体聚集体是早期药物发现中最常见的人工产物，无特异性地隔离和抑制靶蛋白。了解它们的结构和机制对于开发控制甚至偶尔利用这些粒子的工具至关重要。不幸的是，它们的多分散性和瞬态稳定性阻止了对某些基本性质的探索，例如它们如何包装。染料稳定的胶体聚集体表现出增强的均匀性和稳定性相比，传统的胶体聚集体，使调查的一些这些属性。通过小角度X射线散射和多角度光散射，对距离分布函数表明，染料稳定的胶体填充，而不是中空的，球。共配制胶体的稳定性使得能够调查它们对结合DNA、肽或折叠蛋白质的偏好，以及它们从另一个中纯化一个的能力。共配制的胶体显示出很少的结合DNA的能力。相应地，胶体优先从甚至1600倍过量的肽中隔离蛋白质，这些肽本身是相同蛋白质消化的结果。这可能反映了蛋白质在与胶体的表面-表面相互作用中具有的亲合力优势。这是第一次，胶体可以显示出对特定蛋白质的偏好高达90倍。装载到胶体上，结合的酶可以旋转下来，重新悬浮，并释放回缓冲液中，恢复其大部分活性。这些意见的胶体机制和效用的影响将被考虑。
Colloidal aggregates of small molecules are the most common artifact in early drug discovery, sequestering and inhibiting target proteins without specificity. Understanding their structure and mechanism has been crucial to developing tools to control for, and occasionally even exploit, these particles. Unfortunately, their polydispersity and transient stability have prevented exploration of certain elementary properties, such as how they pack. Dye-stabilized colloidal aggregates exhibit enhanced homogeneity and stability when compared to conventional colloidal aggregates, enabling investigation of some of these properties. By small-angle X-ray scattering and multiangle light scattering, pair distance distribution functions suggest that the dye-stabilized colloids are filled, not hollow, spheres. Stability of the coformulated colloids enabled investigation of their preference for binding DNA, peptides, or folded proteins, and their ability to purify one from the other. The coformulated colloids showed little ability to bind DNA. Correspondingly, the colloids preferentially sequestered protein from even a 1600-fold excess of peptides that are themselves the result of a digest of the same protein. This may reflect the avidity advantage that a protein has in a surface-to-surface interaction with the colloids. For the first time, colloids could be shown to have preferences of up to 90-fold for particular proteins over others. Loaded onto the colloids, bound enzyme could be spun down, resuspended, and released back into buffer, regaining most of its activity. Implications of these observations for colloid mechanisms and utility will be considered.