停乳链球菌GapC的免疫特性分析及B细胞抗原表位的筛选与鉴定

Analysis of immune characteristics of Streptococcus dysgalactiae GapC and screening and identification of B cell antigens and epitopes

旨在表达牛乳源停乳链球菌3-磷酸甘油醛脱氢酶(GapC),预测并鉴定其B细胞抗原表位。本研究采用PCR扩增停乳链球菌GapC基因,构建重组质粒p ET-28a-TR-X16087-2。经IPTG诱导表达后纯化GapC蛋白,并以纯化的GapC蛋白免疫小鼠,采用ELISA方法检测Ig G抗体效价及抗体分型,分析对小鼠的免疫保护效力。结果显示,成功表达了大小为44 ku的GapC蛋白,对小鼠的免疫保护率为76%。预测并筛选出3个B细胞抗原表位,鉴定了1个优势抗原表位BP3:^(196)PHRGGDLRRARAGAAN^(206)。上述结果表明,本研究成功表达了停乳链球菌GapC蛋白,对其免疫效果进行了评估,预测与鉴定了其B细胞表位,为GapC蛋白功能和表位疫苗的研究奠定了基础。

The aim of this study was to express the glyceraldehyde-3-phosphate dehydrogenase(GapC)of Streptococcus dysgalactiae isolated from bovine milk and predict,identify its B-cell epitopes.In this study,PCR was used to amplify the GapC gene of Streptococcus dysgalactiae,and recombinant expression plasmid pET-28a-TR-X16087-2 was constructed.Then GapC recombinant protein was induced and expressed.The purified protein was used to immunize mice.The antibody titer and antibody typing of IgG was analyzed using ELISA method.The protection effect was tested with mice.The result showed that GapC protein with a size of 44 ku was successfully expressed.After imuunization,the mice protection rate was 76%.Three B-cell epitopes were predicted and screened,and one dominant epitope BP3 was identified:^(196)PHRGGDLRRARAGAAN^(206).In conclusion,the GapC of S.dysgalactiaewas successfully expressed and its immunogenicity was analyzed.The B-cell epitopes of GapC were predicted and identified.This data laid the foundation for the further study of GapC protein function and epitope vaccine.