Zheng Kai, Laurence Jennifer S, Kuczera Krzysztof, Verkhivker Gennady, Middaugh C Russell, Siahaan Teruna J
Department of Pharmaceutical Chemistry, The University of Kansas, Lawrence, KS 66047, USA.
Chem Biol Drug Des. 2009 Jun;73(6):584-98. doi: 10.1111/j.1747-0285.2009.00818.x.
The objectives of this work were to express the EC5 domain of E-cadherin and determine its structural characteristics as well as to evaluate the binding properties of HAV and BLG4 peptides to EC5 using spectroscopic methods. Homophilic interactions of E-cadherins are responsible for cell-cell adhesion in the adherens junctions of the biological barriers (i.e. intestinal mucosa and blood-brain barriers). The EC5 domain of E-cadherin has an important role in T-cell adhesion to intestinal mucosa via alpha(E)beta(7) integrin-E-cadherin interactions. In this study, the expressed EC5 has a high thermal stability (T(m) = 64.3 degrees C); it also has two stable conformations at room temperature, which convert to one conformation at approximately 54.5 degrees C. NMR and FTIR showed that HAV and BLG4 peptides bind to EC5. HSQC-NMR showed that either Asn or Gln of EC5 was involved in the interactions with HAV and BLG4 peptides. EC5 underwent a conformational change upon interaction with the HAV and BLG4 peptides. Finally, the binding properties of both peptides were modeled by docking experiments, and the results suggest that Asn-46 and Asn-75 of EC5 could be involved during the interaction with the peptides and that the Ser and Trp residues of the HAV and BLG4 peptides, respectively, were important for binding to EC5.
这项工作的目标是表达E-钙黏蛋白的EC5结构域，确定其结构特征，并使用光谱方法评估HAV和BLG4肽与EC5的结合特性。E-钙黏蛋白的同源相互作用负责生物屏障（即肠黏膜和血脑屏障）黏附连接中的细胞间黏附。E-钙黏蛋白的EC5结构域在T细胞通过α(E)β(7)整合素-E-钙黏蛋白相互作用黏附到肠黏膜中起重要作用。在本研究中，表达的EC5具有高热稳定性（熔点Tm = 64.3℃）；它在室温下也有两种稳定构象，在约54.5℃时转变为一种构象。核磁共振（NMR）和傅里叶变换红外光谱（FTIR）表明HAV和BLG4肽与EC5结合。异核单量子相干（HSQC）-NMR表明EC5的天冬酰胺或谷氨酰胺参与了与HAV和BLG4肽的相互作用。EC5与HAV和BLG4肽相互作用时发生了构象变化。最后，通过对接实验对两种肽的结合特性进行了建模，结果表明EC5的天冬酰胺-46和天冬酰胺-75可能参与了与肽的相互作用，并且HAV和BLG4肽的丝氨酸和色氨酸残基分别对与EC5的结合很重要。
