GENES FOR HYDROXYPROLINE-RICH GLYCOPROTEINS

细胞-细胞/细胞-基质识别现象对于许多生物学过程是必不可少的。 生物过程（正常和致病）。 我们希望确定在 详细描述了控制特定大分子缔合的分子规则 参与细胞粘附和细胞外基质组装。 衣藻 是一个很好的实验生物，由于这种研究的设施， 生物化学结构和基因分析 承认/遵守 分子（凝集素）和细胞外基质（细胞壁）成分 从衣原体中分离出来， 富含羟脯氨酸的糖蛋白（HRGP）可能有一个共同的 进化遗产 每类HRGP都参与特异性结合 以模拟其体内功能的方式在体外具有活性。 什么 构成了一个特定的结合域？ 这些领域在以下方面有何不同？ 同源（正和负）凝集素分子？ 都是同源结构域 负责细胞壁组件的自组装？ 是凝集素， 矩阵HRGP的多基因家族的成员？ cDNA文库将是 由每次交配的营养细胞和配子细胞的poly A+ RNA构建 型（mt+和mt-）的衣原体，在表达载体LambdagtII中， 或在Lambdagt 10中。 将使用抗体（LambdagtII）筛选文库 或合成的寡核苷酸（Lambdagt 10）探针，以鉴定编码 对于mt+和mt-凝集素和GPl，晶体层的组分 与凝集素有关的细胞壁的几个 的搜索. 将鉴定凝集素的候选结合序列 差异杂交。 针对相应的抗体产生的抗体 融合蛋白将用于抑制测定和表位作图。 功能域将进一步检查，使用合成肽 抗肽抗体。 GP 1结合结构域，在相同的实施方案中鉴定， 方式，将绘制并与凝集素序列进行比较，以作为证据 的同源性。

Cell-cell/cell-matrix recognition phenomena are essential to numerous biological processes (normal and pathogenic). We wish to determine in detail the molecular rules governing specific macromolecular associations involved in cell-adhesion and extracellular matrix assembly. Chlamydomonas is an excellent experimental organism for such studies due to the facility of biochemical, structural, and genetic analyses. Recognition/adhesion of molecules (agglutinins) and extracellular matrix (cell wall) components have been isolated from Chlamydomonas and shown to be members of a family of hydroxyproline-rich glycoproteins (HRGP's) that may share a common evolutionary heritage. Each class of HRGP engages in specific binding activities in vitro in a manner that mimics their in vivo function. What constitutes a specific binding domain? How do these domains differ between homologous (plus and minus) agglutinin molecules? Are homologous domains responsible for self-assembly of cell wall components? Are agglutinin and matrix HRGP's members of a multigene family? cDNA libraries will be constructed from poly A+ RNA of vegetative and gametic cells of each mating type (mt+ and mt-) of Chlamydomonas, in the expression vector Lambdagtll, or in Lambdagt10. Libraries will be screened using antibody (Lambdagtll) or synthetic oligonucleotide (Lambdagt10) probes to identify inserts coding for mt+ and mt- agglutinins and GPl, a component of the crystalline layer of the cell wall that appears to be related to the agglutinins by several criteria. Candidate binding sequences for agglutinins will be identified by differential hybridization. Antibodies produced against corresponding fusion proteins, will be used for inhibition assays and epitope-mapping. Functional domains will be examined further, using synthetic peptides anti-peptide antibodies. GPl binding domains, identified in the same manner, will be mapped and compared with agglutinin sequences for evidence of homology.