用于蛋白质结晶的形核剂研究综述

The acquisition of protein crystals is a crucial step in the analysis of three-dimensional structure of proteins, the production of medical drugs and the construction of self-assembled nanostructures. For example, when we use X-ray diffraction technology to determine the three-dimensional structure of protein, first of all, we need to obtain the high quality of protein crystals. Protein crystallization has two steps, nucleation and growth. The progress from the undersaturation zone through metastable zone to nucleation zone is called nucleation. When the protein crystal nucleus passes through the nucleation zone to the metastable zone, the crystal nucleus grows and ripening occurs. In the process of protein crystallization, formation of protein crystal nucleus is fundamental. The whole crystallization system has the same probability to nucleate in homogeneous nucleation process. In this process, the protein starts to nucleate in the nucleation zone only when the degree of supersaturation of the protein solution crystallization system could overcome the nucleation barrier. Thus homogeneous nucleation process has certain limitations for the low concentration solution in crystallization. In comparison to homogenous nucleation, with the presence of nucleating agents, the heterogeneous nucleation of the protein crystals faces relatively small resistance and barriers in this process. Hence the use of nucleating agents has critical significance for low-crystallizability proteins or low-initial-concentration crystallization process. With the development of the structural biology, nucleating agents is still a hot issue in crystallization methodology studies. The adsorption effect of porous microspheres is beneficial to the formation of disordered protein molecular clusters, and thus could promote protein nucleation. Adding porous microspheres will lead to both the increment of the number of crystallization screening hits and the quality of the resultant crystals. The use of artificial seed crystal could make protein molecules orderly arranged, and keep a low-concentration condition throughout the crystal nucleation growth process. And the crosslinked crystal seed has greater application potential due to its higher stability. The novel cross-diffusion microbatch plate enables the crystallization system to realize concentration adjusting through a relatively slow cross-diffusion process and to finally reach mutual equilibrium, and in consequence, causes significant increment in the protein crystalline screening hits and the crystal quality improvement, as the number of crystallization screening hits for protein protease K increases from 39 to 47, and the diffraction resolution of crystal increases from 1.66 Å to 1.54 Å. Some properties of the substrate, such as electrostatic effect, hydrophobic interaction and hydrogen bonding, can be also used to promote the protein molecules aggregation and nucleation. This paper summarizes the influence which nucleating agents exert on the protein crystallization from two perspectives of physical and chemical interactions, also roughly discusses the future prospect and research directions.