How phosphorylation of peptides affects their interaction with 14‐3‐3η domains

Abstract

Members of the 14-3-3 domain family have important functions as adapter domains.Via an amphipathic groove on their protein surface they typically bind to disordered C-terminals of other proteins. Importantly, binding partners of 14-3-3 domains usually contain a phosphorylated serine or threonine residue at their binding interface and possess one of three different sequence motifs. Binding of the respective unphosphorylated versions of the peptides is typically strongly disfavored. There is a wealth of structural and thermodynamic data available for the phosphorylated forms but not for the unphosphorylated forms as the binding affinities seem to be too weak to be measurable experimentally. Here, we characterized the mechanistic details that govern the preference for the binding of phosphorylated peptides to 14-3-3η domains by means of molecular dynamics (MD) simulations. We found that the phosphate group is ideally coordinated in the binding pocket whereas the respective unphosphorylated side-chain counterpart is not. Thus, the binding preference results from the tight coordination of the phosphorylated residue at the center of the binding interface. Furthermore, MD simulations of 14-3-3η dimers showed a preference for the simultaneous binding of two phosphorylated peptides in agreement with their experimentally observed cooperativity.

How Peptides Bind to PSD-95/Discs-Large/ZO-1 Domains

“…Representative structures of peptide S (top left) and peptide pS1 (bottom left) bound to the PDZ2 domain. These structures were extracted similar to our previous study by finding the centroid, the frame with the highest sum of similarities, of all trajectory frames of 1 μs long plain MD simulations. Shown on the right are 2D sketches of the binding pocket and the relevant hydrogen bonds between protein and peptide, respectively.…”

“… a Similar to our previous study or using the “double system in a single box” setup at 283 K and n NaCl = 150 mmol. All simulations were performed using a switching time of 1 ns, if not mentioned otherwise.…”

“…One source of error may be the nonavailability of standard state and electrostatic finite-size corrections for “double system in a single box” setups. In order to test if the established correction terms of a different setup improve the results, we also performed alchemical simulations of the bound and free states separately and applied the standard state and electrostatic finite-size corrections similar to our previous study . The resulting binding free energy differences and corrections are listed in Table .…”

“…Here it is expected that some artifacts of changes in charge will cancel out. The second method is to simulate the bound and free states separately and to account for artifacts in the alchemical simulations using the correction scheme described in our previous work based on the correction scheme developed by Rocklin et al Similar corrections schemes have been derived by Reif and Oostenbrink as well as Öhlknecht et al Corrections have to be used for periodicity-induces net-charge interactions and undersolvation, , for residual integrated potential effects , and for discrete solvent effects. , Additionally, an empirical correction term , has to be applied as well. In this project we performed both types of simulations to understand their differences.…”

Structural insights on the effects of mutation of a charged binding pocket residue on phosphopeptide binding to 14‐3‐3ζ protein

“…Their sequence and structures are largely identical and yet their functions are often non redundant 13,14 . Due to the multivariate roles they have in our cell, 14‐3‐3 proteins are widely studied using experimental 15 as well as computational (especially molecular dynamics [MD] simulations 16–18 ) methods to gain understanding about their structure and function. The minor differences in sequence and structure of various isoforms may have a role to play in distinct functions even within the same pathway 19 …”