IL-21 Interacts With Heparin And Initially Reveals The Conformation Of Its Intracellular Domain Of Gamma Receptor Subunits

Interleukin-21 is an I-type cytokine that has an important pleiotropic immunoregulatory effect,it also has an effect on both humoral and T-cell immunity.IL-21 is known to exert its biological function by first binding to the ectodomain of its receptor to form the IL-21/IL-21R?/IL-21Ry ternary complex,then,the signal is transmitted through the transmembrane region of the receptor into the cell,leading to the activation of the JAK/STAT signaling pathway.However,the detailed molecular mechanism of the process remains to be studied.This thesis tries to deepen understanding of this signal transduction process by studying the characteristics of IL-21 and its receptors in the following aspects.(1)Previous experiments showed that IL-21 signal transduction may be related to the microenvironment.That is,HSPG on the cell surface and matrix may affect the conformation of IL-21 and thus affect its biological function.Here we use nuclear magnetic resonance to study specific participating areas.(2)Protein structure and function are related to each other,and the stability of the conformation directly affects its functionality.In the previous period,we selected high stability mutants by computer simulation.Here,we expressed and purified enough samples to explore the stability of the IL-21 mutant conformations by liquid nuclear magnetic resonance.(3)We studied the transmembrane region of the IL-21R? receptor subunit.First,the structure of the transmembrane region of the receptor was explored,and its secondary structure information was initially obtained.Due to its strong hydrophobicity and small molecular weight,the transmembrane region is very challenging for its sample preparation,so we tried several different separation and purification methods.(4)The cell membrane may be involved in signal transduction as an important factor.To study the interaction of the intracellular domain of the alpha receptor subunit with the cell membrane,alapha-subunit protein samples were successfully prepared.The fluorescence spectra and lipid strips experiments were used to identify the interaction between the protein and the plasma membrane in vitro.The intracellular domain protein is not easy to crystallize,the structure is loose,and the stability is low,therefore,it is difficult to use X-ray diffraction and liquid nuclear magnetic means to obtain to three-dimensional structure information.Based on the specificity of the intracellular domain structure of the receptors,we firstly attempted to using chemical cross-linking and mass spectrometry to obtain the three-dimensional structural information of the alapha subunit intracellular domain.