Structural characterization of an intrinsically disordered protein complex using integrated small‐angle neutron scattering and computing

Abstract

Characterizing structural ensembles of intrinsically disordered proteins (IDPs) and intrinsically disordered regions (IDRs) of proteins is essential for studying structure–function relationships. Due to the different neutron scattering lengths of hydrogen and deuterium, selective labeling and contrast matching in small‐angle neutron scattering (SANS) becomes an effective tool to study dynamic structures of disordered systems. However, experimental timescales typically capture measurements averaged over multiple conformations, leaving complex SANS data for disentanglement. We hereby demonstrate an integrated method to elucidate the structural ensemble of a complex formed by two IDRs. We use data from both full contrast and contrast matching with residue‐specific deuterium labeling SANS experiments, microsecond all‐atom molecular dynamics (MD) simulations with four molecular mechanics force fields, and an autoencoder‐based deep learning (DL) algorithm. From our combined approach, we show that selective deuteration provides additional information that helps characterize structural ensembles. We find that among the four force fields, a99SB‐disp and CHARMM36m show the strongest agreement with SANS and NMR experiments. In addition, our DL algorithm not only complements conventional structural analysis methods but also successfully differentiates NMR and MD structures which are indistinguishable on the free energy surface. Lastly, we present an ensemble that describes experimental SANS and NMR data better than MD ensembles generated by one single force field and reveal three clusters of distinct conformations. Our results demonstrate a new integrated approach for characterizing structural ensembles of IDPs.

Impact of Nonnative Interactions on the Binding Kinetics of Intrinsically Disordered p53 with MDM2: Insights from All-Atom Simulation and Markov State Model Analysis

“…The studied systems ranged from short intrinsically disordered peptides with a length of 12−19 amino-acid residues 70 to small intrinsically disordered domains with ∼50 amino-acid residues. 71 The discrepancy between these works may potentially be reconciled by two reasons. First, the CHARMM36m force field is specifically optimized for IDPs when used with the TIP3P model, 72 which explains its superior performance compared to the above-mentioned protein force fields.…”

“…They together provide good agreement with CD, NMR, and SANS results. The studied systems ranged from short intrinsically disordered peptides with a length of 12–19 amino-acid residues to small intrinsically disordered domains with ∼50 amino-acid residues . The discrepancy between these works may potentially be reconciled by two reasons.…”

A science-driven approach to optimize the design for a biological small-angle neutron scattering instrument

“…Smallangle neutron scattering (SANS) is an important part of this tool kit and has been used to investigate a wide variety of systems that range from isolated and purified biomolecules in solution (Vandavasi et al 2016;Brady et al, 2022) to complex systems including plant and soil architectures (Phyo et al, 2017;Pingali et al, 2017;Yang et al, 2022), and even whole cells (Tehei et al, 2007). The uniqueness of the structural information that comes from SANS measurements arises from the ability to exert precise control over the scattering contrast by deuteration, making it possible to probe specific parts of complex and dynamic assemblies in a complex environment (Liberton et al, 2013;Stingaciu et al, 2016;Sparks et al, 2018;Bush et al, 2019;Pingali et al, 2020;Qian et al, 2020;Chen et al, 2023;Gurumoorthy et al, 2023).…”