Kinoshita Masanao, Yamaguchi Shimpei, Matsumori Nobuaki
Department of Chemistry, Graduate School of Science, Kyushu University, Fukuoka, 819-0395, Japan.
Sci Rep. 2020 Dec 21;10(1):22188. doi: 10.1038/s41598-020-79083-7.
Ordered/disordered phase separation occurring in bio-membranes has piqued researchers' interest because these ordered domains, called lipid rafts, regulate important biological functions. The structure of the ordered domain has been examined with artificial membranes, which undergo macroscopic ordered/disordered phase separation. However, owing to technical difficulties, the local structure inside ordered domains remains unknown. In this study, we employed electron diffraction to examine the packing structure of the lipid carbon chains in the ordered domain. First, we prepared dehydrated monolayer samples using a rapid-freezing and sublimation protocol, which attenuates the shrinkage of the chain-packing lattice in the dehydration process. Then, we optimised the electron flux to minimise beam damage to the monolayer sample. Finally, we developed low-flux scanning electron diffraction and assessed the chain packing structure inside the ordered domain formed in a distearoylphosphatidylcholine/dioleoylphosphatidylcholine binary monolayer. Consequently, we discovered that the ordered domain contains multiple subdomains with different crystallographic axes. Moreover, the size of the subdomain is larger in the domain centre than that near the phase boundary. To our knowledge, this is the first study to reveal the chain packing structures inside an ordered domain.
生物膜中发生的有序/无序相分离引起了研究人员的兴趣，因为这些被称为脂筏的有序结构域调节着重要的生物学功能。有序结构域的结构已通过人工膜进行了研究，人工膜会发生宏观的有序/无序相分离。然而，由于技术困难，有序结构域内部的局部结构仍然未知。在本研究中，我们采用电子衍射来研究有序结构域中脂质碳链的堆积结构。首先，我们使用快速冷冻和升华方案制备脱水单层样品，该方案可减轻脱水过程中链堆积晶格的收缩。然后，我们优化了电子通量，以尽量减少对单层样品的束损伤。最后，我们开发了低通量扫描电子衍射，并评估了在二硬脂酰磷脂酰胆碱/二油酰磷脂酰胆碱二元单层中形成的有序结构域内的链堆积结构。因此，我们发现有序结构域包含多个具有不同晶轴的子结构域。此外，子结构域的尺寸在结构域中心比在相界附近更大。据我们所知，这是第一项揭示有序结构域内部链堆积结构的研究。
