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 "Ceff calculator" (http://ceffapp.chemeslab.org), which will allow user-friendly prediction of experimentally inaccessible parameters.
Keywords: Affinity enhancement; Autoinhibition; Avidity; C(eff) calculator; Disordered linker; Effective concentration; Worm-like chain.
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