Hemorrhagic bacillus coli of intestine 0157:H7 shiga toxin IIB epitope peptide and uses thereof
A kind of Escherichia coli, enterohemorrhagic technology, applied in the field of medicine and biology
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Embodiment 1
[0035] Example 1 Screening and identification of B epitopes:
[0036] 1. Prediction and synthesis of epitope peptides
[0037] Retrieve the Stx2 of EHEC O157:H7 in the NCBI structural database and the crystal structure analysis diagram of Stx2 and ligand adenine (No. 1R4P and 2GA4), website: http: / / www.ncbi.nlm.nih.gov / sites / entrez? db=Structure). Based on the three-dimensional structure of the interaction between Stx2 and the ligand adenine, and based on the prediction principles of B epitope peptides exposed on the surface of the protein, containing stretches and random coils or β-turns, and peptides with a length of about 15-30 amino acids, the selected Amino acid peptides on the toxic fragment of Stx2A1, the predicted amino acid peptides were analyzed using DNAStar software for hydrophilicity (Kyte-Doolittle scheme), surface accessibility (Emini scheme), plasticity (Karp lus-Schulz scheme), secondary structure ( Garnier and Chou-Fasman scheme) and antigenicity (Jame...
Embodiment 2
[0056] Example 2 Screening of Protective B Cell Epitope Peptides by Immune Mice Challenge Test
[0057] Female Balb / c mice aged 6-8 weeks were randomly divided into 5 groups, 5 mice in each group, and 4 B cell epitope peptides (P1-KLH, P2-KLH, P3-KLH, P4- KLH) subcutaneous multi-point plus intraperitoneal injection to immunize Balb / c mice, each injected with 0.51ml, containing 100μg of antigen, and the immunization time was 0, 2, 4, 6, and 8 weeks, a total of 5 times, the first time adding Freund's complete Adjuvant Add Freund's incomplete adjuvant for the 2nd to 4th times, and intraperitoneal injection of the same dose without adjuvant for the last immunization. Blood was collected 7 days after the 2nd, 3rd, 4th, and 5th immunizations, and the immune Balb was detected by indirect ELISA. / c mouse serum titer (operating steps as before); calculate antibody positivity conversion rate, mean geometric titer (GMT) and average growth multiple. Ten days after the last immunization...
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