Refinement of α-Synuclein Ensembles Against SAXS Data: Comparison of Force Fields and Methods.

Mustapha Carab Ahmed,Alexander Jussupow,Line K Skaanning,Estella A Newcombe,Carlo Camilloni,Annette E Langkilde,Kresten Lindorff-Larsen,Birthe B Kragelund

doi:10.3389/fmolb.2021.654333

Abstract

The inherent flexibility of intrinsically disordered proteins (IDPs) makes it difficult to interpret experimental data using structural models. On the other hand, molecular dynamics simulations of IDPs often suffer from force-field inaccuracies, and long simulation times or enhanced sampling methods are needed to obtain converged ensembles. Here, we apply metainference and Bayesian/Maximum Entropy reweighting approaches to integrate prior knowledge of the system with experimental data, while also dealing with various sources of errors and the inherent conformational heterogeneity of IDPs. We have measured new SAXS data on the protein α-synuclein, and integrate this with simulations performed using different force fields. We find that if the force field gives rise to ensembles that are much more compact than what is implied by the SAXS data it is difficult to recover a reasonable ensemble. On the other hand, we show that when the simulated ensemble is reasonable, we can obtain an ensemble that is consistent with the SAXS data, but also with NMR diffusion and paramagnetic relaxation enhancement data.

Highlights

Disordered Proteins (IDPs) play important roles in a wide range of biological processes including cell signaling and regulation (Uversky et al, 2005; Das et al, 2015; Snead and Eliezer, 2019), and their malfunction or aggregation is linked to neurodegenerative diseases such as Alzheimer’s and Parkinson’s disease
We find that the inclusion of a small angle Xray scattering (SAXS)-restraint in the M&M simulation resulted in the generation of a reliable and heterogenous conformational ensemble that improved the agreement with the nuclear magnetic resonance (NMR)
Our result provide insight into how and when experimental SAXS data can be used to refine ensembles of Intrinsically Disordered Proteins (IDPs), and the role played by the force field as a ‘prior’ in these Bayesian/Maximum entropy approaches

Summary

Introduction

Disordered Proteins (IDPs) play important roles in a wide range of biological processes including cell signaling and regulation (Uversky et al, 2005; Das et al, 2015; Snead and Eliezer, 2019), and their malfunction or aggregation is linked to neurodegenerative diseases such as Alzheimer’s and Parkinson’s disease. Due to the dynamic nature of IDPs and their inherent conformational heterogeneity, IDPs are not amenable to high-resolution characterization solely through experimental measurements To characterize their structural and dynamic properties it is often necessary to integrate various biophysical experiments, and nuclear magnetic resonance (NMR). Refinement of Ensembles Against SAXS Data spectroscopy (Dyson and Wright, 2001), small angle Xray scattering (SAXS) (Bernado and Svergun, 2012), circular dichroism (Chemes et al, 2012), and single-molecule Förster resonance energy transfer (sm-FRET) (LeBlanc et al, 2018) have been widely used to characterize the structural properties of IDPs. For instance, pulsed-field-gradient NMR diffusion and SAXS experiments are especially useful to quantify the level of compaction of the IDP.

Methods

Results

Conclusion