STRUCTURE-FUNCTION OF TRANSFORMING GROWTH FACTOR BETA

多功能多肽转化生长因子β（TGF-β） β），是蛋白水解衍生自一个较大的 前体分子先前的研究表明， 一种被称为β-潜伏期相关肽（beta-latency-associated peptide，beta-Leptide）的蛋白质， 与较小的二聚体成熟生长因子以非共价方式结合。 这 紧密和特异性的相互作用完全是由于β-环糊精， 在加热和酸处理时变性;热和酸基本上 对成熟生长因子无影响。 本报告的总体目标 一项建议是确定TGF-β与 他们的β-LAPs和研究潜在的激活机制。 这些研究的结果对于理解 TGF-β生物活性的调节。 去检查每一个测试- LAP，我们将在转染的CHO组织培养细胞中表达它们 独立于成熟的TGF-β，允许产生大量的 一定量的生物功能性β-内酰胺酶。 这种约束力的来源 蛋白质将允许研究测量的解离常数 成熟TGF-β和用于生产多克隆和单克隆 抗体的 抗体将允许研究评估结构 与其他β-LAP相关。 因为他们基本上没有 抗体对不同的β-LAP反应，这些免疫球蛋白 将提供新的免疫试剂福特方法， 免疫沉淀或组织分布研究。 此外该 不同的表达的β-LAPs将使用已经纯化的 建立了纯化方案，并用于测试这些复合物是否 与细胞表面或细胞外基质结合。 最后， 从TGF-β 2a和TGF-β 2b，和TGF-β 3将表达为 嵌合体与β-内酰胺酶，以研究必要的结构区域， 延迟。 这些研究应该加强我们对潜在的 复杂的，并提供线索的调节控制的TGF-β。

The multi-functional polypeptides, transforming growth factor betas (TGF- betas), are proteolytically derived from the carboxyl-terminus of a larger precursor molecule. Previous studies indicate that a dimer of the pro piece, known as beta-latency-associated peptide (beta-LAP), binds in a noncovalent manner with the smaller, dimeric, mature growth factor. This tight and specific interaction is entirely due to beta-LAP, being denatured upon heating and acid treatment; heat and acid have essentially no effects on the mature growth factor. The overall objective of this proposal is to define the molecular interaction of the TGF-betas with their beta-LAPs and to investigate potential activation mechanisms. Results from these studies are important for the understanding of the regulation of TGF-beta biological activity. To examine each of the beta- LAPs, we will express them in transfected CHO tissue culture cells independent of mature TGF-beta, allowing for the production of large amounts of a biologically, functional beta-LAP. This source of binding protein will allow for studies measuring the dissociation constants of mature TGF-beta and for the production of both polyclonal and monoclonal antibodies. Antibodies will permit studies to assess the structural relatedness with other beta-LAPs. Since their are essentially no antibodies reacting toward the diverse beta-LAPs, these immunoglobulins will provide new immunereagents ford approaches such as immuneprecipitation or tissue distribution studies. In addition, the different, expressed beta-LAPs will be purified, using an already established purification scheme, and used to test whether these complexes bind to cell surfaces or to extracellular matrix. Finally, the beta-LAPs from TGF-beta2a and TGF-beta2b, and TGF-beta3 will be expressed as chimeras with the beta-LAP to investigate structural regions necessary for latency. These studies should enhance our understanding of the latent complex and provide clues to the regulatory controls of the TGF-betas.