Institut für Angewandte Physik, Universität Tübingen , Auf der Morgenstelle 10, 72076 Tübingen, Germany.
J Am Chem Soc. 2015 Feb 4;137(4):1485-91. doi: 10.1021/ja510533x. Epub 2015 Jan 23.
We present a real-time study of protein crystallization of bovine β-lactoglobulin in the presence of CdCl(2) using small-angle X-ray scattering and optical microscopy. From observing the crystallization kinetics, we propose the following multistep crystallization mechanism that is consistent with our data. In the first step, an intermediate phase is formed, followed by the nucleation of crystals within the intermediate phase. During this period, the number of crystals increases with time, but the crystal growth is slowed down by the surrounding dense intermediate phase due to the low mobility. In the next step, the intermediate phase is consumed by nucleation and slow growth, and the crystals are exposed to the dilute phase. In this stage, the number of crystals becomes nearly constant, whereas the crystals grow rapidly due to access to the free protein molecules in the dilute phase. This real-time study not only provides evidence for a two-step nucleation process for protein crystallization but also elucidates the role and the structural signature of the metastable intermediate phase in this process.
我们使用小角 X 射线散射和光学显微镜实时研究了牛β-乳球蛋白在 CdCl2 存在下的结晶过程。通过观察结晶动力学，我们提出了以下多步结晶机制，该机制与我们的数据一致。在第一步中，形成了一个中间相，然后在中间相中形成晶体核。在此期间，晶体的数量随时间增加，但由于中间相密度高导致晶体的迁移率低，晶体生长受到抑制。在下一步中，中间相通过成核和缓慢生长被消耗，晶体暴露在稀相中。在这个阶段，晶体的数量几乎保持不变，而晶体由于可以进入稀相中游离的蛋白质分子而快速生长。这项实时研究不仅为蛋白质结晶的两步成核过程提供了证据，还阐明了亚稳中间相在该过程中的作用和结构特征。
