FUNCTIONAL ANTIGENS OF TRICHINELLA SPIRALIS

两种针对日本血吸虫L1幼虫的保护性诱导抗原 旋毛虫的结构将被研究 并利用免疫学、生物化学和分子生物学技术 生物学方法。表达和基因组文库的构建 合适的载体将被构建和克隆DNA分子 编码48K和50/55K蛋白的基因将被分离。 阳性克隆的表达将通过克隆鉴定 多价兔抗48K和50/55K抗体的免疫检测 抗体。含有表位的融合肽将被测试 它们在小鼠体内诱导保护的能力。Southern杂交分析 将决定编码两种基因的DNA的基因组排列 抗原。旋毛虫各期的Northern分析 (L1-L4幼虫、成虫和新生幼虫)将揭示阶段- 每个抗原的基因表达的特异性。按顺序排列 将确定每个抗原的cDNA和基因组DNA。 这两种蛋白质都可以被磷酸化，此外，48K 抗原结合二价阳离子，尤其是钙离子。因此，多肽 将对每个分子进行作图和测序研究 来确定这两种蛋白质的磷酸化位置， 以及48K分子与钙结合的数量和位置。 初级氨基酸共有序列的计算机分析 和DNA序列将确定疏水性区域和 与其他已知序列的同源性程度。特例 注意调控蛋白与48K之间的相似性 并将给予50/55K抗原，因为初步数据表明 它们可能参与了寄主厌食症的调节 糖酵解。

Two protection-inducing antigens specific to the L1 larva of Trichinella spiralis will be investigated regarding their structure and function using immunological, biochemical and molecular biological approaches. Expression and genomic libraries in suitable vectors will be constructed and cloned DNA molecules encoding both the 48K and 50/55K protein will be isolated. Expression of positive cDNA clones will be identified by colony immune assay with polyvalent rabbit anti-48K and 50/55K antibodies. Fusion peptides containing epitopes will be tested for their ability to induce protection in mice. Southern blot analysis will determine the genomic arrangement of DNA encoding both antigens. Northern analysis on each stage of Trichinella spiralis (L1-L4 larva, adult and newborn larva) will reveal the stage- specificity of gene expression for each antigen. The sequence of cDNA and genomic DNA for each antigen will be determined. Both proteins can be phosphorylated, and in addition, the 48K antigen binds divalent cations, particularly Ca2+. Thus peptide mapping and sequencing studies on each molecule will be carried out to determine the sites of phosphorylation for both proteins, and the number and sites of Ca2+ binding for the 48K molecule. Computer analysis of the primary amino acid consensus sequence and DNA sequence will determine areas of hydrophobicity and degrees of homology with other known sequences. Particular attention to similarities between regulatory proteins and the 48K and 50/55K antigens will be given, since preliminary data indicate that they may be involved in regulation of host anerobic glycolysis.