Recombinant production and characterization of the human mast cell tryptases gamma and delta

类胰蛋白酶是肥大细胞中最丰富和最特异的蛋白质之一。在人类中已经发现了四种类胰蛋白酶，但只有类胰蛋白酶α和β在功能和结构上得到了很好的表征，这表明β是参与肥大细胞与疾病相关的不良功能和最近发现的维持健康的良好功能的关键介质。相比之下，类胰蛋白酶γ和δ几乎没有受到关注，在它们的功能和对肥大细胞生物学的贡献方面一直是孤立的。在这个项目中，我们在酵母和哺乳动物细胞中表达了人类胰蛋白酶γ和δ及其变异体，从而使我们实验室中所有的人类胰蛋白酶都可用。我们目前正在使用这些重组类胰蛋白酶来研究它们的酶活性和调节，阐明它们的结构，并产生它们的检测和抑制工具。我们计划专门使用这些工具来探索膜结合类胰蛋白酶γ的不同特征及其对类胰蛋白酶的中介作用的贡献。我们认为，了解和分离单个类胰蛋白酶的性质和功能对于作为一个整体理解类胰酶以及以任何类胰酶为靶点的治疗干预是至关重要的。

Tryptases are serine proteinases that are among the most abundant and specific proteins of mast cells. Four tryptases have been identified in man, but only tryptase α and β have been functionally and structurally well characterized, showing that β is a key mediator involved in both the disease-related ‘bad’ and the more recently discovered healthmaintaining ‘good’ functions of mast cells. In contrast, tryptase γ and δ have received little attention and have remained orphan in respect to their functions and their contributions to mast cell biology. In this project we have expressed human tryptase γ and δ and variants in yeast and mammalian cells, thereby making all human tryptases available in our lab. We are currently using these recombinant tryptases to study their enzymatic activity and regulation, to elucidate their structures and to generate tools for their detection and inhibition. We plan to employ these tools specifically to explore the distinguishing features of the membrane-bound tryptase γ and its contributions to the mediator actions of tryptases. We believe that understanding and separating the properties and functions of the individual tryptases is essential for the understanding of tryptases as a group and for therapeutic interventions using any tryptase as target.