Glaeser R M
Lawrence Berkeley National Laboratory, University of California Berkeley, Berkeley, California 94720, USA.
J Struct Biol. 1999 Dec 1;128(1):3-14. doi: 10.1006/jsbi.1999.4172.
From a modest beginning with negatively stained samples of the helical T4 bacteriophage tail, electron crystallography has emerged as a powerful tool in structural biology. High-resolution density maps, interpretable in terms of an atomic structure, can be obtained from specimens prepared as well-ordered, two-dimensional crystals, and the resolution achieved with helical specimens and icosahedral viruses is approaching the same goal. A hybrid approach to determining the molecular structure of complex biological assemblies is generating great interest, in which high-resolution structures that have been determined for individual protein components are fitted into a lower resolution envelope of the large complex. With this as background, how much more can be anticipated for the future? Considerable scope still remains to improve the quality of electron microscope images. Automation of data acquisition and data processing, together with the emergence of computational speeds of 10(12) floating point operations per second or higher, will make it possible to extend high-resolution structure determination into the realm of single-particle microscopy. As a result, computational alignment of single particles, i.e., the formation of "virtual crystals," can begin to replace biochemical crystallization. Since single-particle microscopy may remain limited to "large" structures of 200 to 300 kDa or more, however, smaller proteins will continue to be studied as helical assemblies or as two-dimensional crystals. The further development of electron crystallography is thus likely to turn increasingly to the use of single particles and small regions of ordered assemblies, emphasizing more and more the potential for faster, higher throughput.
从对螺旋状T4噬菌体尾部负染样本的初步研究开始，电子晶体学已成为结构生物学中的一种强大工具。通过制备成排列有序的二维晶体的标本，可以获得可根据原子结构进行解释的高分辨率密度图，并且螺旋状标本和二十面体病毒所达到的分辨率也正在接近这一目标。一种用于确定复杂生物组装体分子结构的混合方法正引起人们极大的兴趣，即在这种方法中，已为单个蛋白质组分确定的高分辨率结构被拟合到大型复合物的较低分辨率包络中。以此为背景，未来还能期待多少呢？提高电子显微镜图像质量仍有相当大的空间。数据采集和数据处理的自动化，以及每秒10¹²次或更高的浮点运算计算速度的出现，将使高分辨率结构测定扩展到单颗粒显微镜领域成为可能。结果，单颗粒的计算对齐，即“虚拟晶体”的形成，可能开始取代生化结晶。然而，由于单颗粒显微镜可能仍局限于200至300 kDa或更大的“大型”结构，较小的蛋白质将继续作为螺旋组装体或二维晶体进行研究。因此，电子晶体学的进一步发展可能会越来越多地转向使用单颗粒和有序组装体的小区域，越来越强调更快、更高通量的潜力。
