Developing a molecular dynamics force field for both folded and disordered protein states

Molecular dynamics (MD) simulation is a valuable tool for characterizing the structural dynamics of folded proteins and should be similarly applicable to disordered proteins and proteins with both folded and disordered regions. It has been unclear, however, whether any physical model (force field) used in MD simulations accurately describes both folded and disordered proteins. Here, we select a benchmark set of 21 systems, including folded and disordered proteins, simulate these systems with six state-of-the-art force fields, and compare the results to over 9,000 available experimental data points. We find that none of the tested force fields simultaneously provided accurate descriptions of folded proteins, of the dimensions of disordered proteins, and of the secondary structure propensities of disordered proteins. Guided by simulation results on a subset of our benchmark, however, we modified parameters of one force field, achieving excellent agreement with experiment for disordered proteins, while maintaining state-of-the-art accuracy for folded proteins. The resulting force field, a99SB-disp, should thus greatly expand the range of biological systems amenable to MD simulation. A similar approach could be taken to improve other force fields

Thermal Adaptation of Conformational Dynamics in Ribonuclease H

The relationship between inherent internal conformational processes and enzymatic activity or thermodynamic stability of proteins has proven difficult to characterize. The study of homologous proteins with differing thermostabilities offers an especially useful approach for understanding the functional aspects of conformational dynamics. In particular, ribonuclease HI (RNase H), an 18 kD globular protein that hydrolyzes the RNA strand of RNA:DNA hybrid substrates, has been extensively studied by NMR spectroscopy to characterize the differences in dynamics between homologs from the mesophilic organism E. coli and the thermophilic organism T. thermophilus. Herein, molecular dynamics simulations are reported for five homologous RNase H proteins of varying thermostabilities and enzymatic activities from organisms of markedly different preferred growth temperatures. For the E. coli and T. thermophilus proteins, strong agreement is obtained between simulated and experimental values for NMR order parameters and for dynamically averaged chemical shifts, suggesting that these simulations can be a productive platform for predicting the effects of individual amino acid residues on dynamic behavior. Analyses of the simulations reveal that a single residue differentiates between two different and otherwise conserved dynamic processes in a region of the protein known to form part of the substrate-binding interface. Additional key residues within these two categories are identified through the temperature-dependence of these conformational processes

International Ocean Discovery Program Expedition 393 Preliminary Report South Atlantic Transect 2

The South Atlantic Transect (SAT) is a multidisciplinary scientific ocean drilling experiment designed to investigate the evolution of the oceanic crust and overlying sediments across the western flank of the Mid-Atlantic Ridge. This project comprises four International Ocean Discovery Program expeditions: fully staffed Expeditions 390 and 393 (April–August 2022) built on engineering preparations during Expeditions 390C and 395E that took place without science parties during the height of the Coronavirus Disease 2019 (COVID-19) pandemic. Through operations along a crustal flow line at ~31°S, the SAT recovered complete sedimentary sections and the upper ~40–340 m of the underlying ocean crust formed at a slow to intermediate spreading rate at the Mid-Atlantic Ridge over the past ~61 My. The sediments along this transect were originally spot cored more than 50 y ago during Deep Sea Drilling Project Leg 3 (December 1968–January 1969) to help verify the theories of seafloor spreading and plate tectonics. The SAT expeditions targeted six primary sites on 7, 15, 31, 49, and 61 Ma ocean crust that fill critical gaps in our sampling of intact in situ ocean crust with regards to crustal age, spreading rate, and sediment thickness. Drilling these sites was required to investigate the history, duration, and intensity of the low-temperature hydrothermal interactions between the aging ocean crust and the evolving South Atlantic Ocean. This knowledge will improve the quantification of past hydrothermal contributions to global biogeochemical cycles and help develop a predictive understanding of the impacts of variable hydrothermal processes and exchanges. Samples from the transect of the previously unexplored sediment- and basalt-hosted deep biosphere beneath the South Atlantic Gyre are essential to refine global biomass estimates and examine microbial ecosystems’ responses to variable conditions in a low-energy gyre and aging ocean crust. The transect is located near World Ocean Circulation Experiment Line A10, which provides a baseline for records of carbonate chemistry and deepwater mass properties across the western South Atlantic through key Cenozoic intervals of elevated atmospheric CO2 and rapid climate change. Reconstruction of the history of the deep western boundary current and deepwater formation in the Atlantic basins will yield crucial data to test hypotheses regarding the role of evolving thermohaline circulation patterns in climate change and the effects of tectonic gateways and climate on ocean acidification. During engineering Expeditions 390C and 395E, a single hole was cored through the sediment cover and into the uppermost rocks of the ocean crust with the advanced piston corer (APC) and extended core barrel (XCB) systems at five of the six primary proposed SAT sites. Reentry systems with casing were then installed either into basement or within 10 m of basement at each of those five sites. Expedition 390 (7 April–7 June 2022) conducted operations at three of the SAT sites, recovering 700 m of core (77%) over 30.3 days of on-site operations. Sediment coring, basement coring, and wireline logging were conducted at two sites on 61 Ma crust (Sites U1556 and U1557), and sediment coring was completed at the 7 Ma Site U1559. Expedition 393 operated at four sites, drilling in 12 holes to complete this initial phase of the SAT. Complete sedimentary sections were collected at Sites U1558, U1583, and U1560 on 49, 31, and 15 Ma crust, respectively, and together with 257.7 m of sediments cored during earlier operations, more than 600 m of sediments was characterized. The uppermost ocean crust was drilled at Sites U1558, U1560, and U1583 with good penetration (~130 to ~204 meters subbasement), but at the youngest ~7 Ma Site U1559, only ~43 m of basement penetration was achieved in this initial attempt. Geophysical wireline logs were aquired at Sites U1583 and U1560. Expeditions 390 and 393 established legacy sites available for future deepening and downhole basement hydrothermal and microbiological experiments at Sites U1557, U1560, and U1559 on 61, 15, and 7 Ma crust, respectively

A practical guide to the simultaneous determination of protein structure and dynamics using metainference

Accurate protein structural ensembles can be determined with metainference, a Bayesian inference method that integrates experimental information with prior knowledge of the system and deals with all sources of uncertainty and errors as well as with system heterogeneity. Furthermore, metainference can be implemented using the metadynamics approach, which enables the computational study of complex biological systems requiring extensive conformational sampling. In this chapter, we provide a step-by-step guide to perform and analyse metadynamic metainference simulations using the ISDB module of the open-source PLUMED library, as well as a series of practical tips to avoid common mistakes. Specifically, we will guide the reader in the process of learning how to model the structural ensemble of a small disordered peptide by combining state-of-the-art molecular mechanics force fields with nuclear magnetic resonance data, including chemical shifts, scalar couplings and residual dipolar couplings.Comment: 49 pages, 9 figure

Clustering Heterogeneous Conformational Ensembles of Intrinsically Disordered Proteins with t-Distributed Stochastic Neighbor Embedding

Intrinsically disordered proteins (IDPs) populate a range of conformations that are best described by a heterogeneous ensemble. Grouping an IDP ensemble into “structurally similar” clusters for visualization, interpretation, and analysis purposes is a much-desired but formidable task, as the conformational space of IDPs is inherently high-dimensional and reduction techniques often result in ambiguous classifications. Here, we employ the t-distributed stochastic neighbor embedding (t-SNE) technique to generate homogeneous clusters of IDP conformations from the full heterogeneous ensemble. We illustrate the utility of t-SNE by clustering conformations of two disordered proteins, Aβ42, and α-synuclein, in their APO states and when bound to small molecule ligands. Our results shed light on ordered substates within disordered ensembles and provide structural and mechanistic insights into binding modes that confer specificity and affinity in IDP ligand binding. t-SNE projections preserve the local neighborhood information, provide interpretable visualizations of the conformational heterogeneity within each ensemble, and enable the quantification of cluster populations and their relative shifts upon ligand binding. Our approach provides a new framework for detailed investigations of the thermodynamics and kinetics of IDP ligand binding and will aid rational drug design for IDPs

Interpreting Protein Structural Dynamics from NMR Chemical Shifts

In this investigation, semiempirical NMR chemical shift prediction methods are used to evaluate the dynamically averaged values of backbone chemical shifts obtained from unbiased molecular dynamics (MD) simulations of proteins. MD-averaged chemical shift predictions generally improve agreement with experimental values when compared to predictions made from static X-ray structures. Improved chemical shift predictions result from population-weighted sampling of multiple conformational states and from sampling smaller fluctuations within conformational basins. Improved chemical shift predictions also result from discrete changes to conformations observed in X-ray structures, which may result from crystal contacts, and are not always reflective of conformational dynamics in solution. Chemical shifts are sensitive reporters of fluctuations in backbone and side chain torsional angles, and averaged ¹H chemical shifts are particularly sensitive reporters of fluctuations in aromatic ring positions and geometries of hydrogen bonds. In addition, poor predictions of MD-averaged chemical shifts can identify spurious conformations and motions observed in MD simulations that may result from force field deficiencies or insufficient sampling and can also suggest subsets of conformational space that are more consistent with experimental data. These results suggest that the analysis of dynamically averaged NMR chemical shifts from MD simulations can serve as a powerful approach for characterizing protein motions in atomistic detail

Available kinetic measurements for RNase H homologs.

<p>Kinetics data measured under various conditions for soRNH, ecRNH, and ttRNH.</p