Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark; The Danish Research Institute for Translational Neuroscience (DANDRITE), Aarhus, Denmark; Center for Proteins in Memory (PROMEMO), Aarhus, Denmark.
Instituto de Investigaciones Biotecnológicas "Dr. Rodolfo A. Ugalde", IIB-UNSAM, IIBIO-CONICET, Universidad Nacional de San Martín, San Martín, Argentina.
Methods Enzymol. 2021;647:145-171. doi: 10.1016/bs.mie.2020.09.012. Epub 2020 Nov 13.
Linkers are crucial to the functions of multidomain proteins as they couple functional units to encode regulation such as auto-inhibition, enzyme targeting or tuning of interaction strength. A linker changes reactions from bimolecular to unimolecular, and the equilibrium and kinetics is thus determined by the properties of the linker rather than concentrations. We present a theoretical workflow for estimating the functional consequences of tethering by a linker. We discuss how to: (1) Identify flexible linkers from sequence. (2) Model the end-to-end distance distribution for a flexible linker using a worm-like chain. (3) Estimate the effective concentration of a ligand tethered by a flexible linker. (4) Calculate the decrease in binding affinity caused by auto-inhibition. (5) Calculate the expected avidity enhancement of a bivalent interaction from effective concentration. The worm-like chain modeling is available through a web application called the "C calculator" (http://ceffapp.chemeslab.org), which will allow user-friendly prediction of experimentally inaccessible parameters.
连接子对于多功能蛋白的功能至关重要，因为它们将功能单元连接起来，以编码调节，如自动抑制、酶靶向或相互作用强度的调谐。连接子将反应从双分子转变为单分子，因此平衡和动力学由连接子的性质决定，而不是浓度。我们提出了一种用于估计连接子连接功能后果的理论工作流程。我们讨论了如何：（1）从序列中识别柔性连接子；（2）使用蠕虫链模型化柔性连接子的末端到末端距离分布；（3）估计通过柔性连接子连接的配体的有效浓度；（4）计算自动抑制引起的结合亲和力下降；（5）从有效浓度计算二价相互作用的预期亲和增强。蠕虫链建模可通过称为“C 计算器”的网络应用程序（http://ceffapp.chemeslab.org）获得，该应用程序将允许用户友好地预测实验不可及的参数。
