Khaleel Zinah Hilal, No Young Hyun, Kim Nam Hyeong, Bae Do Hyun, Wu Yibing, Kim Suhyeon, Choi Hojae, Lee Da Eun, Jeong Se Yun, Ko Yoon-Joo, Kim Seong-Gi, Suh Minah, Kim Jin-Chul, DeGrado William F, Kim Ki Hyun, Kim Yong Ho
SKKU Advanced Institute of Nanotechnology (SAINT), Sungkyunkwan University, Suwon 16419, Republic of Korea.
Center for Neuroscience Imaging Research, Institute for Basic Science, Suwon 16419, Republic of Korea.
Proc Natl Acad Sci U S A. 2025 Feb 4;122(5):e2411316122. doi: 10.1073/pnas.2411316122. Epub 2025 Jan 30.
The design of organic-peptide hybrids has the potential to combine our vast knowledge of protein design with small molecule engineering to create hybrid structures with complex functions. Here, we describe the computational design of a photoswitchable Ca-binding organic-peptide hybrid. The designed molecule, designated Ca-binding switch (CaBS), combines an EF-hand motif from classical Ca-binding proteins such as calmodulin with a photoswitchable group that can be reversibly isomerized between a spiropyran (SP) and merocyanine (MC) state in response to different wavelengths of light. The MC/SP group acts both as a photoswitch as well as an optical sensor of Ca binding. Photoconversion of the SP to the corresponding MC unmasks an acidic phenol, which CaBS uses as an integral part of both its Ca-binding site as well as its tertiary and quaternary structure. By design, the SP state of CaBS is monomeric, while the Ca-bound form of the MC state is an obligate dimer, with two Ca-binding sites formed at the interface of a domain-swapped dimer. Thus, light and Ca were expected to serve as an "AND gate" that powers a change in backbone structure/dynamics, oligomerization state, and fluorescence properties of the designed molecule. CaBS was designed using Rosetta and molecular dynamics simulations, and experimentally characterized by nuclear magnetic resonance, isothermal titration calorimetry, and optical titrations. These data illustrate the potential of combining small molecule engineering with de novo protein design to develop sensors whose conformation, association state, and optical properties respond to multiple environmental cues.
有机肽杂化物的设计有潜力将我们在蛋白质设计方面的丰富知识与小分子工程相结合，以创造具有复杂功能的杂化结构。在此，我们描述了一种光开关钙结合有机肽杂化物的计算设计。所设计的分子称为钙结合开关（CaBS），它将来自经典钙结合蛋白（如钙调蛋白）的EF手基序与一个光开关基团相结合，该基团可响应不同波长的光在螺吡喃（SP）和部花青（MC）状态之间可逆异构化。MC/SP基团既作为光开关，又作为钙结合的光学传感器。SP向相应MC的光转换会暴露一个酸性酚，CaBS将其用作钙结合位点以及三级和四级结构的一个组成部分。通过设计，CaBS的SP状态是单体，而MC状态的钙结合形式是一个必然的二聚体，在结构域交换二聚体的界面处形成两个钙结合位点。因此，光和钙有望作为一个“与门”，驱动所设计分子的主链结构/动力学、寡聚化状态和荧光特性发生变化。CaBS是使用Rosetta和分子动力学模拟设计的，并通过核磁共振、等温滴定量热法和光学滴定进行了实验表征。这些数据说明了将小分子工程与从头蛋白质设计相结合以开发其构象、缔合状态和光学性质对多种环境线索作出响应的传感器的潜力。
