ENGINEERING PROTEINS WITH UNUSUAL ARCHITECTURES

本申请将侧重于方法论的发展和 具有特殊结构的蛋白质的治疗应用。已选择 我们设计的蛋白质将被循环和分支蛋白质。目标 循环蛋白包括白细胞介素2受体拮抗剂，它是 目前正在进行临床试验，作为一种降低脓毒症严重程度的药物 关节炎；监测肽，一种胆囊收缩素释放因子和 可能对治疗消化系统疾病有用；防御素是 一种具有抗艾滋病相关活性的广谱抗生素 病原体。目标分支蛋白将包括疟疾疫苗 含有来自裂殖子表面蛋白的保护性抗原 (MSP-1)。这种抗原是迄今为止最有希望的候选疫苗。这个 分支设计将MSP-1抗原的多聚体拷贝整合到 小肽核心基质为多抗原肽(MAP)。这个 MSP-1来源的抗原是构象依赖的，含有多个 二硫键。加上T辅助表位和内置佐剂 组装成地图，它将提供一种完整的疫苗 明确的结构。这种疟疾疫苗将进行保护试验。 Carol Long博士在小鼠模型中提出的抗感染挑战 疟疾方面的顶尖专家。含有HIV-1抗原的MAP的一个版本是 目前处于第一阶段临床试验，作为候选艾滋病疫苗。 这个应用的统一主题是开发和应用 我们实验室最近构思的结构域连接策略。 它是一种将完全不受保护的多肽连接或循环的方法 蛋白质片段通过多肽键而不被激活。这种方法是 非常适合于合成环状和支链蛋白 不能通过重组DNA方法直接访问，并且是 用常规的多肽合成方法难以获得。这个 结构域连接策略是一种有机和多肽相结合的方法 用于治疗应用的工程蛋白质的化学。

This application will focus on the methodological development and therapeutic applications of proteins with unusual architectures. Selected proteins for our design will be circularized and branch proteins. Target circularized proteins include interleukin-l receptor antagonist which is currently in clinical trials as a drug to reduce severity of sepsis and arthritis; monitor peptide which is a cholecystokinin-releasing factor and may be useful for treatment of digestive disorders; and defensin which is a broad-spectrum antibiotic with promising activity against AIDS-related pathogens. Target branch proteins will include a malaria vaccine containing the protective antigen derived from merozoite surface protein (MSP-1). This antigen is the most promising vaccine candidate to date. The branch design will incorporate multimeric copies of MSP-1 antigen in a small peptidyl core matrix as multiple antigen peptides (MAPs). The antigen derived from MSP-1 is conformation-dependent and contains multiple disulfide bonds. Together with a T-helper epitope and a built-in adjuvant assembled to form MAPs, it will provide a complete vaccine with an unambiguous structure. This malaria vaccine will be tested for protection against infection challenge in the mouse model by Dr. Carol Long who is a leading expert in malaria. A version of MAPs containing a HIV-1 antigen is currently in Phase I clinical trials as a candidate AIDS vaccine. The unifying theme of this application is the development and application of the domain ligation strategy recently conceptualized in our laboratory. It is a method to link or circularize totally unprotected peptide and protein segments via a peptide bond without activation. This method is well suited for the synthesis of circularized and branch proteins which are inaccessible directly by the recombination DNA method and are difficult to obtain by the conventional peptide synthesis approaches. The domain ligation strategy is a combined approach of organic and peptide chemistry in engineering proteins for therapeutic applications.