Kinases play central roles in signaling cascades, relaying information from the outside to the inside of mammalian cells. De novo designed protein switches capable of interfacing with tyrosine kinase signaling pathways would open new avenues for controlling cellular behavior, but, so far, no such systems have been described. Here we describe the de novo design of two classes of protein switch that link phosphorylation by tyrosine and serine kinases to protein-protein association. In the first class, protein-protein association is required for phosphorylation by the kinase, while in the second class, kinase activity drives protein-protein association. We design systems that couple protein binding to kinase activity on the immunoreceptor tyrosine-based activation motif central to T-cell signaling, and kinase activity to reconstitution of green fluorescent protein fluorescence from fragments and the inhibition of the protease calpain. The designed switches are reversible and function in vitro and in cells with up to 40-fold activation of switching by phosphorylation.

激酶在信号级联中发挥核心作用，将信息从哺乳动物细胞的外部传递到内部。从头设计的能够与酪氨酸激酶信号通路连接的蛋白质开关将为控制细胞行为开辟新途径，但到目前为止，还没有描述过这样的系统。在这里，我们描述了两类蛋白质开关的从头设计，它们将酪氨酸和丝氨酸激酶的磷酸化与蛋白质-蛋白质关联联系起来。在第一类中，激酶磷酸化需要蛋白质-蛋白质关联，而在第二类中，激酶活性驱动蛋白质-蛋白质关联。我们设计的系统将蛋白质结合与 T 细胞信号传导中枢的免疫受体酪氨酸激活基序上的激酶活性结合起来，并将激酶活性与片段中绿色荧光蛋白荧光的重建和蛋白酶钙蛋白酶的抑制结合起来。设计的开关是可逆的，并且在体外和细胞中发挥作用，通过磷酸化将开关激活高达 40 倍。