Department of Biochemistry, University of Washington, Seattle, WA, USA.
Institute for Protein Design, University of Washington, Seattle, WA, USA.
Nature. 2019 Aug;572(7768):205-210. doi: 10.1038/s41586-019-1432-8. Epub 2019 Jul 24.
Allosteric regulation of protein function is widespread in biology, but is challenging for de novo protein design as it requires the explicit design of multiple states with comparable free energies. Here we explore the possibility of designing switchable protein systems de novo, through the modulation of competing inter- and intramolecular interactions. We design a static, five-helix 'cage' with a single interface that can interact either intramolecularly with a terminal 'latch' helix or intermolecularly with a peptide 'key'. Encoded on the latch are functional motifs for binding, degradation or nuclear export that function only when the key displaces the latch from the cage. We describe orthogonal cage-key systems that function in vitro, in yeast and in mammalian cells with up to 40-fold activation of function by key. The ability to design switchable protein functions that are controlled by induced conformational change is a milestone for de novo protein design, and opens up new avenues for synthetic biology and cell engineering.
蛋白质功能的变构调节在生物学中广泛存在,但对于从头设计蛋白质来说具有挑战性,因为它需要明确设计具有可比自由能的多个状态。在这里,我们通过调节竞争的分子内和分子间相互作用来探索从头设计可切换蛋白质系统的可能性。我们设计了一个具有单个界面的静态五螺旋“笼”,该界面可以与末端“闩锁”螺旋分子内相互作用,或者与肽“键”分子间相互作用。在闩锁上编码了用于结合、降解或核输出的功能基序,只有当键从笼中置换出闩锁时,这些功能基序才起作用。我们描述了正交笼键系统,该系统在体外、酵母和哺乳动物细胞中均起作用,通过键的诱导构象变化可将功能激活高达 40 倍。能够设计受诱导构象变化控制的可切换蛋白质功能是从头设计蛋白质的一个里程碑,并为合成生物学和细胞工程开辟了新途径。