Molecular analysis of thymopentin binding to HLA-DR molecules.
Thymopentin (TP5) triggers an immune response by contacting with T cells; however the molecular basis of how TP5 achieves this process remains incompletely understood. According to the main idea of immunomodulation, we suppose that it would be necessary for TP5 to form complex with human class II major histocompatibility complex DR molecules (HLA-DR) before TP5 interacts with T cells. The uptake of TP5 by EBV-transformed B cells expressing HLA-DR molecules and the histogram of fluorescence intensities were observed by using fluorescent- labeled TP5, testifying the direct binding of TP5 to HLA-DR. The binding specificity was confirmed by the inhibition with unlabeled TP5, suggesting the recognition of TP5 by HLA-DR. To confirm the interaction between TP5 and HLA-DR, the complex formation was predicted by using various modeling strategies including six groups of trials with different parameters, alanine substitutions of TP5, and the mutants of HLA-DR. The results demonstrated that TP5 and its alanine substitutions assumed distinct conformations when they bound to HLA-DR. The observation further showed that there was flexibility in how the peptide bound within the binding cleft. Also, the molecular analysis supplemented a newly important discovery to the effect of Val anchor on TP5 binding HLA-DR, and revealed the important effects of Glu11 and Asn62 on the recognition of TP5. These results demonstrated the capability of TP5 to associate with HLA-DR in living antigen presenting cells (APC), thereby providing a new and promising strategy to understand the immunomodulation mechanism induced by TP5 and to design potential immunoregulatory polypeptides.
Liu
Zuojia
Z
State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun, Jilin, China. jin.wang.1@stonybrook.edu
Zheng
Xiliang
X
Wang
Jin
J
Wang
Erkang
E
D001402:B-Lymphocytes / Q000378:metabolism; D001667:Binding, Competitive; D002461:Cell Line, Transformed; D016650:Fluorescein-5-isothiocyanate; D006684:HLA-DR Antigens / Q000378:metabolism*; D004854:Herpesvirus 4, Human / Q000378:metabolism; D006801:Humans; D011485:Protein Binding; D016305:Thymopentin / Q000378:metabolism*
D006684:HLA-DR Antigens; D016650:Fluorescein-5-isothiocyanate; D016305:Thymopentin