Encounter and Binding of Camp at the Binding Domain of Mlok1

Ligand-protein binding processes play an essential role in biological systems, be it signalling immune response or enzymatic activity. An experimentally well-studied system is the binding of cyclic adenosine monophosphate (cAMP) at the cyclic nucleotide binding domain (CNBD) of the bacterial potassium channel MloK1. The channel's conductivity is modulated by cAMP binding and is prototypical for cyclic nucleotide gated ion channels.Two models were postulated to describe the binding process: The first is a two-step model consisting of bulk-surface diffusion and surfing-binding site rolling. The second consists of a diffusion process into a binding funnel combined with stochastic barrier crossing.Here we present a comprehensive molecular dynamics study of the primary cAMP binding events, i.e. the ligands path to the binding site in the CNBD, to identify substates, establish a model of the ligand pathways and to predict an effective on-rate for the binding process. Available experimental high precision measurements of the binding kinetics render the system suitable for comparison.We identified multiple surface substates that play a crucial role for the dynamics for high ligand concentrations. Using rate estimates from the MD simulations and extrapolation to low, experimentally accessible ligand concentrations we ruled out the two-step models. We quantified the conformational confinement and the probability of binding barrier crossing to obtain values for the funnel model. The overall estimate for the on-rate rate constant was estimated to be 30/(µs∗mol/l), which is in excellent agreement with the experimental measurements.