Cloning and expression of the major 47-kilodalton surface immunogen of Treponema pallidum in Escherichia coli.
Cloning and expression of the major 47-kilodalton surface immunogen of Treponema pallidum in Escherichia coli.
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梅毒螺旋体主要 47 千道尔顿表面免疫原在大肠杆菌中的克隆和表达。
DOI：
10.1128/iai.54.2.500-506.1986
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发表时间：
1986
影响因子：
3.1
通讯作者：
Goldberg,MS
中科院分区：
文献类型：
--
作者：
Norgard,MV;Chamberlain,NR;Swancutt,MA;Goldberg,MS
针对47千道尔顿（kDa）的主要外膜表面免疫原的毒性梅毒螺旋体亚种的单克隆抗体。pallidum用于选择表达47-kDa免疫原的大肠杆菌重组克隆。克隆的表型依赖于宿主细胞中重组质粒的存在。Southern杂交结果表明，克隆的T.苍白球亚种苍白球DNA序列是T.苍白球亚种苍白球基因组DNA排列。从实验感染T.苍白球亚种梅毒和人继发梅毒血清与克隆特异性反应，而正常人血清或来自正常兔血清的免疫球蛋白G不与克隆反应。Southern杂交的结果表明，同源的47-kDa的免疫原基因是不存在的至少四种非致病性密螺旋体测试，以及从总兔基因组DNA。兔抗T.噬菌体生物型Reiter（非病原体密螺旋体）抗血清和针对常见密螺旋体决定簇的单克隆抗体与克隆不反应。免疫印迹和放射免疫沉淀实验表明，重组大肠杆菌（E。coli）和天然（T.苍白球亚种苍白球）形式的抗原具有相同的电泳迁移率。重组47-kDa免疫原的可用性提供了一个新的机会，生化分析的蛋白质，结构-功能研究，检查其在微生物发病机制中的作用，并评估其诊断和疫苗的潜力。
Monoclonal antibodies directed against the 47-kilodalton (kDa) major outer membrane surface immunogen of virulent Treponema pallidum subsp. pallidum were used to select Escherichia coli recombinant clones expressing the 47-kDa immunogen. The phenotype of the clones was dependent on the presence of recombinant plasmid in the host cell. Southern hybridization revealed that the cloned T. pallidum subsp. pallidum DNA sequence was an accurate representation of the T. pallidum subsp. pallidum genomic DNA arrangement. Purified immunoglobulin G from rabbits experimentally infected with T. pallidum subsp. pallidum and human secondary syphilitic sera specifically reacted with the clones, while normal human serum or immunoglobulin G from normal rabbit serum did not. Results of Southern hybridization indicated that a homologous 47-kDa immunogen gene was absent in at least four species of nonpathogenic treponemes tested, as well as from total rabbit genomic DNA. Rabbit anti-T. phagedenis biotype Reiter (treponemal nonpathogen) antiserum and a monoclonal antibody directed against a common treponemal determinant were unreactive with the clones. Western blotting and radioimmunoprecipitation experiments with specific monoclonal antibodies revealed that the recombinant (E. coli) and native (T. pallidum subsp. pallidum) forms of the antigen had identical electrophoretic mobilities. The availability of recombinant 47-kDa immunogen provides a new opportunity for biochemical analysis of the protein, structure-function studies, examination of its role in microbial pathogenesis, and assessment of its diagnostic and vaccinogenic potentials.