FUNCTIONAL ANTIGENS OF TRICHINELLA SPIRALIS

两种特异性保护性诱导抗原 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 生物学方法。 表达和基因组文库 将构建合适的载体并克隆DNA分子 将分离编码48 K和50/55 K蛋白的蛋白。 将通过菌落鉴定阳性cDNA克隆的表达 多价兔抗48 K和50/55 K免疫测定 抗体的 将检测含有表位的融合肽的 它们在小鼠中诱导保护的能力。 Southern印迹分析 将决定编码这两种基因的DNA的基因组排列 抗原 旋毛虫各虫态的北方分析 （L1-L4幼虫，成虫和新生幼虫）将揭示阶段- 每种抗原的基因表达特异性。 的序列 将测定每种抗原的cDNA和基因组DNA。 这两种蛋白质都可以被磷酸化，此外，48 K 抗原结合二价阳离子，特别是Ca 2+。 因此肽 将对每个分子进行定位和测序研究， 为了确定两种蛋白质的磷酸化位点， 以及48 K分子与Ca ~（2+）结合的数目和位点。 一级氨基酸共有序列的计算机分析 DNA序列将决定疏水性区域， 与其他已知序列的同源性程度。 特别 注意调节蛋白和48 K之间的相似性 将给予50/55 K抗原，因为初步数据表明， 它们可能参与调节宿主的厌氧 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.