Siebert H C, von der Lieth C W, Kaptein R, Beintema J J, Dijkstra K, van Nuland N, Soedjanaatmadja U M, Rice A, Vliegenthart J F, Wright C S, Gabius H J
Institut für Physiologische Chemie, Tierärztliche Fakultät, Ludwig-Maximilians-Universität, München, Germany.
Proteins. 1997 Jun;28(2):268-84.
Carbohydrate recognition by lectins often involves the side chains of tyrosine, tryptophan, and histidine residues. These moieties are able to produce chemically induced dynamic nuclear polarization (CIDNP) signals after laser irradiation in the presence of a suitable radical pair-generating dye. Elicitation of such a response in proteins implies accessibility of the respective groups to the light-absorbing dye. In principle, this technique is suitable to monitor surface properties of a receptor and the effect of ligand binding if CIDNP-reactive amino acids are affected. The application of this method in glycosciences can provide insights into the protein-carbohydrate interaction process, as illustrated in this initial study. It focuses on a series of N-acetylglucosamine-binding plant lectins of increasing structural complexity (hevein, pseudohevein, Urtica dioica agglutinin and wheat germ agglutinin and its domain B), for which structural NMR- or X-ray crystallographic data permit a decision of the validity of the CIDNP method-derived conclusions. On the other hand, the CIDNP data presented in this study can be used for a rating of our molecular models of hevein, pseudohevein, and domain B obtained by various modeling techniques. Experimentally, the shape and intensity of CIDNP signals are determined in the absence and in the presence of specific glycoligands. When the carbohydrate ligand is bound, CIDNP signals of side chain protons of tyrosine, tryptophan, or histidine residues are altered, for example, they are broadened and of reduced intensity or disappear completely. In the case of UDA, the appearance of a new tryptophan signal upon ligand binding was interpreted as an indication for a conformational change of the corresponding indole ring. Therefore, CIDNP represents a suitable tool to study protein-carbohydrate interactions in solution, complementing methods such as X-ray crystallography, high-resolution multidimensional nuclear magnetic resonance, transferred nuclear Overhauser effect experiments, and molecular modeling.
凝集素对碳水化合物的识别通常涉及酪氨酸、色氨酸和组氨酸残基的侧链。在存在合适的自由基对生成染料的情况下，激光照射后，这些部分能够产生化学诱导动态核极化（CIDNP）信号。蛋白质中引发这种反应意味着相应基团可接触到吸光染料。原则上，如果CIDNP反应性氨基酸受到影响，该技术适用于监测受体的表面性质以及配体结合的效果。如本初步研究所展示的，此方法在糖科学中的应用能够深入了解蛋白质 - 碳水化合物相互作用过程。它聚焦于一系列结构复杂度递增的N - 乙酰葡糖胺结合植物凝集素（橡胶素、类橡胶素、荨麻凝集素和麦胚凝集素及其结构域B），对于这些凝集素，结构核磁共振或X射线晶体学数据能够判定由CIDNP方法得出的结论是否有效。另一方面，本研究中呈现的CIDNP数据可用于对通过各种建模技术获得的橡胶素、类橡胶素和结构域B的分子模型进行评估。在实验中，CIDNP信号的形状和强度在不存在和存在特定糖配体的情况下被测定。当碳水化合物配体结合时，酪氨酸、色氨酸或组氨酸残基侧链质子的CIDNP信号会发生改变，例如，它们会变宽、强度降低或完全消失。在荨麻凝集素的情况下，配体结合时新色氨酸信号的出现被解释为相应吲哚环构象变化的一个指示。因此，CIDNP是研究溶液中蛋白质 - 碳水化合物相互作用的合适工具，可补充诸如X射线晶体学、高分辨率多维核磁共振、转移核Overhauser效应实验和分子建模等方法。
