Mouse HCV (hepatitis C virus) polypeptide epitope combined with MHC-I molecule and application thereof
A hepatitis virus, MHC-I technology, applied in the direction of viral peptides, medical preparations containing active ingredients, applications, etc., can solve problems such as research that has not been reported in the literature, and achieve convenient clinical application, large commercial industrialization value, The effect of improving accuracy
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Embodiment 1
[0033] The screening method of embodiment 1 polypeptide epitope
[0034] The polypeptide sequence obtained in the present invention is directly retrieved from the peptide library through the method of combining the supermotif, quantitative motif and artificial neural network scheme, and maintains the binding property with MHC-I molecules.
[0035] 1. Acquisition of amino acid sequence of murine hepatitis III virus S protein
[0036] The full-length amino acid sequence (330 amino acids in total) of the natural mouse hepatitis III virus S protein found from the international open shared gene bank NCBI GeneBank is expressed as: SEQ NO.1.
[0037] 2. Acquisition of supermotifs predicted by polypeptide epitopes
[0038] Supermotifs are peptide motifs based on identical and similar anchor residues for the acceptance of antigenic peptides in the same leukocyte antigen (HLA) family, or even HLA allotypes of different families. Easy with H-2K K In the combined 8-peptide sequence, th...
Embodiment 2
[0047] Example 2 Three-dimensional structure analysis and molecular dynamics simulation analysis of polypeptide epitope binding to MHC-I
[0048] Use the Silicon graphics workstation and Insight II software to establish the predicted peptide and H-2K epitopes for the above-mentioned obtained peptide epitopes. K Combined three-dimensional structure and molecular dynamics simulation, including the following methods: ①Molecular model construction, using the Discover3.0 module in the InsightII software package (using CVFF force field), the octapeptide from the S protein of mouse hepatitis III virus and H-2K K Compounds of molecules are subjected to molecular dynamics simulations. H-2K K The initial coordinates come from the protein structure database (PDB number 1ZT1), and each octapeptide is obtained by using the Bioploymer module to perform amino acid substitutions on the polypeptide molecules in 1ZT1. The simulation process is as follows: First, the H-2K K Fixed with β2m, t...
Embodiment 3
[0051] Example 3 Obtaining of the polypeptide epitope of the present invention
[0052] (1) Synthesis of peptides
[0053] The standard Fmoc protocol is used for the polypeptide, and the arginine is coupled twice, and the peptide chain is extended from the carboxy-terminus to the amino-terminus according to the polypeptide sequence. After the peptide is synthesized, the corresponding cleavage enzyme is selected for cleavage, and the protective group of the peptide is removed at the same time to obtain the crude peptide.
[0054] (2) Purification and molecular weight analysis of polypeptide epitopes
[0055] The polypeptide epitopes obtained above were purified and analyzed by RP-HPLC, and the purity of the four synthesized polypeptides determined by RP-HPLC was above 95%. The results of mass spectrometry showed that the measured values of the relative molecular masses of the peptides were basically consistent with the theoretical values, which could be used for subsequent ...
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