Macromolecular Crystallography Research with Synchrotron Radiation

在这一年中，我们已经证明了硒脲的实际用途，这种试剂可以浸泡在蛋白质的天然晶体中，提供强烈的硒异常衍射信号，用于解决大分子的新晶体结构。硒脲是我们在去年向生物晶体学界提出的一种简单而强大的载体，用于同步加速器x射线光束线的相位衍射数据测量。该试剂使我们能够解决XXX的晶体结构，XXX是植物中氨基酸组氨酸生物合成途径中的一种酶。在方法学方面，我们对储存在蛋白质数据库（PDB）中的蛋白质晶体结构中出现的不同类型的非结晶螺旋轴进行了分析。在与Wlodawer博士（Frederick MCL）和其他同事的合作下，对各种小分子配体蛋白质复合物的分析进行了彻底的分析和验证，经常纠正文献和PDB中可用结果的原始次优解释。分析了Z-DNA的异常晶体结构，并将其解释为高度假对称和六重单面孪晶标本，这是以前未观察到的这些类型的晶体。在解决和解释具有生物学或医学重要性的蛋白质晶体结构的活动中，我们从氨基酸丝氨酸和组氨酸以及脂肪多胺的生物合成途径中选择了模式植物酶，这些酶不是在动物中合成的，因此可能与开发对抗各种病原体的活性剂有关，在药理学或农业中具有潜在的应用前景。豆科植物紫花苜蓿的组氨酸途径包含7种酶，可催化多种反应（有些是双功能的）。所有这些都被表达出来，五个晶体结构被解决，解释和最近发表。其余的结晶化了，对它们的研究还在继续。从丝氨酸途径中获得了四种酶并进行了结晶，并对其中两种酶的结构进行了完整的表征和发表。近年来研究了多胺（腐胺精胺、亚精胺和热精胺）生物合成过程中几种酶的晶体结构。对热精胺合成酶结构的解释已发表，对其他氨基丙基转移酶的研究仍在继续。

During this year we have shown the practical usefulness of selenourea, the reagent that can be soaked into the native crystals of proteins to provide the strong anomalous diffraction signal of selenium used for solving novel crystal structures of macromolecules. Selenourea was in the previous year proposed to the biocrystallographic community by us as a simple and powerful vehicle for phasing diffraction data measured at synchrotron X-ray beam lines. This reagent allowed us to solve the crystal structure of XXX, one of the enzymes from the biosynthetic pathway of the amino acid histidine in plants. Within the methodological aspects of our activities, we conducted the analysis of non-crystallographic screw axes of different types occurring in protein crystal structures stored in the Protein Data Bank (PDB). In collaboration with Dr. Wlodawer (MCL in Frederick) and other colleagues the analysis various complexes of proteins with small molecular ligands was thoroughly analyzed and validated, often correcting the original suboptimal interpretation of results available in the literature and in the PDB. The unusual crystal structures of Z-DNA were analyzed, and interpreted as highly pseudosymmetric and six-fold merohedrally twinned specimens, not observed previously for these types of crystals. Within our activity directed at solving and interpreting crystal structures of proteins having biological or medical importance, we worked on selected model plant enzymes from the biosynthetic pathways of amino acids serine and histidine as well as aliphatic polyamines, which are not synthesized in animals and therefore may be relevant for development of agents active against various pathogens, with potential application in pharmacology or agriculture. The histidine pathway in the legume plant Medicago truncatula contain seven enzymes, catalyzing several reactions (some are bifunctional). All of them are expressed and five crystal structures are solved, interpreted and recently published. The rest are crystallized and the work on them continues. Four enzymes from the serine pathway are obtained and crystallized and structures of two of them fully characterized and published. The crystal structures of several enzymes involved in biosynthesis of polyamines putrescine spermine, spermidine and thermospermine were recently solved. The interpretation of the structure of thermospermine synthetase is published and work on other aminopropyltransferases continues.