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首页> 外文期刊>Journal of Molecular Biology >What is the average conformation of bacteriophage T4 lysozyme in solution? A domain orientation study using dipolar couplings measured by solution NMR.
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What is the average conformation of bacteriophage T4 lysozyme in solution? A domain orientation study using dipolar couplings measured by solution NMR.

机译：溶液中噬菌体T4溶菌酶的平均构象是什么？使用通过溶液NMR测量的偶极偶合进行畴取向研究。

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摘要

Lysozyme from T4 bacteriophage is comprised of two domains that are both involved in binding substrate. Although wild-type lysozyme has been exclusively crystallized in a closed form that is similar to the peptidoglycan-bound conformation, a more open structure is thought to be required for ligand binding. To determine the relative arrangement of domains within T4 lysozyme in the solution state, dipolar couplings were measured in several different dilute liquid crystalline media by solution NMR methods. The dipolar coupling data were analyzed with a domain orientation procedure described previously that utilizes high- resolution X-ray structures. The cleft between the domains is significantly larger in the average solution structure than what is observed in the X-ray structure of the ligand-free form of the protein (approximately 17 degrees closure from solution to X-ray structures). A comparison of the solution domain orientation with X-ray-derived structures in the protein data base shows that the solution structure resembles a crystal structure obtained for the M6I mutant. Dipolar couplings were also measured on the lysozyme mutant T21C/T142C, which was oxidized to form an inter-domain disulfide bond (T4SS). In this case, the inter-domain solution structure was found to be more closed than was observed in the crystal (approximately 11 degrees). Direct refinement of lysozyme crystal structures with the measured dipolar couplings using the program CNS, establishes that this degree of closure can be accommodated whilst maintaining the inter-domain cystine bond. The differences between the average solution conformations obtained using dipolar couplings and the crystal conformations for both forms of lysozyme investigated in this study illustrate the impact that crystal packing interactions can have on the arrangement of domains within proteins and the importance of alternative methods to X-ray crystallography for evaluating inter-domain structure. Copyright 2001 Academic Press.
机译：来自T4噬菌体的溶菌酶由两个都与结合底物有关的结构域组成。尽管野生型溶菌酶已经完全以类似于肽聚糖结合的构象的闭合形式结晶，但是认为配体结合需要更开放的结构。为了确定溶液状态下T4溶菌酶中域的相对排列，通过溶液NMR方法在几种不同的稀液晶介质中测量了偶极偶合。通过使用高分辨率X射线结构的先前描述的畴取向程序分析了偶极耦合数据。结构域之间的裂缝在平均溶液结构中比在无配体形式的蛋白质的X射线结构中观察到的裂隙大得多（从溶液到X射线结构大约17度闭合）。蛋白质数据库中溶液结构域方向与X射线衍生结构的比较表明，溶液结构类似于为M6I突变体获得的晶体结构。还对溶菌酶突变体T21C / T142C进行了偶极偶合，该突变体被氧化形成域间二硫键（T4SS）。在这种情况下，发现域间溶液结构比晶体中观察到的更紧密（约11度）。使用程序CNS用测得的偶极偶合对溶菌酶晶体结构进行直接精制，可以确定在保持域间胱氨酸键的同时，可以适应这种闭合程度。使用偶极偶合获得的平均溶液构象与两种溶菌酶的晶体构象之间的差异，说明了晶体堆积相互作用可能对蛋白质中结构域的排列产生影响，以及X射线替代方法的重要性晶体学评估域间结构。版权所有2001学术出版社。

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