摘要
为了获得滑液囊支原体(Mycoplasma synoviae,MS)二氢硫辛酰胺脱氢酶(PdhD)的蛋白表达产物,分析蛋白的免疫原性,采用DNAStar Protean软件分析PdhD的二级结构和主要抗原域。将PdhD基因的第593、731、1712、1833 nt处的A突变为G,使改造成功的基因能够在大肠杆菌BL21中正确表达;经IPTG诱导后获得约70 ku的蛋白。对PdhD蛋白的622个氨基酸进行二级结构的预测,Garnier-Robson法预测出18个α螺旋中心、12个β折叠区段和17个T转角区段,Chou-Fasman法预测出24个α螺旋中心、23个β折叠区段和37个T转角区段,Karplus-Schulz法预测出42个柔性区域。PdhD的主要抗原域为Gln27-Phe36、Vla38-Val51、Ala89-Ala99、Pro137-Asp159、Gly307-Ile336、Gly369-Gly386、Gly397-Gln416、Ala433-Val447、Ieu486-Tyr504、Ala541-Asn549。免疫原性分析发现,PdhD重组蛋白能与兔源Anti-MS血清发生强烈反应,与兔源Anti-MG血清和兔源血清Blank均不发生反应。PdhD具有免疫原性,可作为候选抗原用于亚单位疫苗的研发。
In order to obtain the protein expression product of Mycoplasma synoviae(MS)dihydrolipoamide dehydrogenase(PdhD)and analyze its immunogenicity,DNAStar Protean software was used to analyze the secondary structure and main antigen domain of Pd⁃hD.A at positions 593,731,1712,and 1833 nt of the PdhD gene was mutated to G,so that the successfully modified gene could be expressed correctly in Escherichia coli BL21;after induction with IPTG,a protein of approximately 70 ku was obtained.The secondary structure of PdhD protein was predicted for 622 amino acids.The Garnier Robson method predicted 18 alpha helix centers,12 beta folding segments,and 17 T-corner segments.The Chou Fasman method predicted 24 alpha helix centers,23 beta folding segments,and 37 T-corner segments.The Karplus Schulz method predicted 42 flexible regions.The main antigenic domain of PdhD was Gln27-Phe36,Vla38-Val51,Ala89-Ala99,Pro137-Asp159,Gly307-Ile336,Gly369-Gly386,Gly397-Gln416,Ala433-Val447,Ieu486-Tyr504,and Ala541-Asn549.Immunogenicity analysis revealed that PdhD recombinant protein could strongly react with rab⁃bit-derived Anti-MS serum,but did not react with rabbit-derived Anti-MG serum or rabbit-derived serum Blank.PdhD had immuno⁃genicity and could be used as a candidate antigen for the development of subunit vaccines.
作者
曹晓怡
胡巧
张文婷
邓兰兰
郭云清
卢琴
张蓉蓉
张鹤平
罗青平
CAO Xiao-yi;HU Qiao;ZHANG Wen-ting;DENG Lan-lan;GUO Yun-qing;LU Qin;ZHANG Rong-rong;ZHANG He-ping;LUO Qing-ping(School of Life Sciences and Food Engineering,Hebei University of Engineering,Handan 056038,Hebei,China;Institute of Animal Husbandry and Veterinary Medicine,Hubei Academy of Agricultural Sciences,Wuhan 430064,China;Hubei Hongshan Laboratory,Wuhan 430070,China)
出处
《湖北农业科学》
2024年第12期205-210,共6页
Hubei Agricultural Sciences
基金
河北省重点研发计划项目乡村振兴技术创新专项(22327311D)
湖北省农业科学院青年科学基金项目(2024NKYJJ19)
国家肉鸡产业技术体系项目(CARS-41)
湖北省重点研发计划项目(2022BBA0055)
湖北省科技重大专项(2020ABA016)
湖北省自然科学基金重点类项目(2021CFA019)。
