Modeling the native ensemble of PhuS using enhanced sampling MD and HDX-ensemble reweighting

Kihn, Kyle C.; Wilson, Tyree; Smith, Aleah; Bradshaw, Richard T.; Wintrode, Patrick L.; Forrest, Lucy R.; Wilks, Angela; Deredge, Daniel

doi:10.1016/j.bpj.2021.11.010

Cited by 12 publications

(15 citation statements)

References 49 publications

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“…In general, structural fluctuations over ps to ns timescales may be accessible to conventional, unbiased MD. Motions across µs to ms timescales, for example the refolding of local structural features in a Heme binding protein [55], are likely to require enhanced sampling approaches [27] (although note that motions only seen on even longer timescales may be associated with global unfolding and EX1-type kinetics, which is as yet unsuitable for HDXer analysis). When using enhanced sampling techniques, users should endeavor to include structures that are representative of the equilibrium dynamics within the candidate ensemble, and no others.…”

Section: Curating a Structural Ensemblementioning

confidence: 99%

Interpreting hydrogen-deuterium exchange experiments with molecular simulations: Tutorials and applications of the HDXer ensemble reweighting software [Article v1.0]

et al. 2022

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Hydrogen-deuterium exchange (HDX) is a comprehensive yet detailed probe of protein structure and dynamics and, coupled to mass spectrometry, has become a powerful tool for investigating an increasingly large array of systems. Computer simulations are often used to help rationalize experimental observations of exchange, but interpretations have frequently been limited to simple, subjective correlations between microscopic dynamical fluctuations and the observed macroscopic exchange behavior. With this in mind, we previously developed the HDX ensemble reweighting approach and associated software, HDXer, to aid the objective interpretation of HDX data using molecular simulations. HDXer has two main functions; first, to compute H-D exchange rates that describe each structure in a candidate ensemble of protein structures, for example from molecular simulations, and second, to objectively reweight the conformational populations present in a candidate ensemble to conform to experimental exchange data. In this article, we first describe the HDXer approach, theory, and implementation. We then guide users through a suite of tutorials that demonstrate the practical aspects of preparing experimental data, computing HDX levels from molecular simulations, and performing ensemble reweighting analyses. Finally we provide a practical discussion of the capabilities and limitations of the HDXer methods including recommendations for a user's own analyses. Overall, this article is intended to provide an up-to-date, pedagogical counterpart to the software, which is freely available at https://github.com/Lucy-Forrest-Lab/HDXer.

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Section: Curating a Structural Ensemblementioning

confidence: 99%

Interpreting hydrogen-deuterium exchange experiments with molecular simulations: Tutorials and applications of the HDXer ensemble reweighting software [Article v1.0]

et al. 2022

Self Cite

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“…Next, we evaluated the models with the HDXer software using the percent deuterium uptake values from our HDX-MS experiments as input ( 49, 50 ) (Fig. 4B, S12A-B, Table S3) .…”

Section: Resultsmentioning

confidence: 99%

“…The ten nsp7-11 output models were assessed against the experimental (i) HDX-MS, (ii) XL-MS and the (iii) SAXS data: (i) Agreement of models to HDX-MS data was completed using HDXer which generated theoretical deuterium uptake values for the models to compare to experimental values ( 49, 50 ). Smaller RMSE indicates better agreement of models to experimental data.…”

Section: Methodsmentioning

confidence: 99%

Biochemical and structural insights into SARS-CoV-2 polyprotein processing by Mpro

Yadav

Courouble

Dey

et al. 2022

Preprint

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SARS-CoV-2, a human coronavirus, is the causative agent of the COVID-19 pandemic. Its ~30 kb RNA genome is translated into two large polyproteins subsequently cleaved by viral papain-like protease and main protease (Mpro/nsp5). Polyprotein processing is essential yet incompletely understood. We studied Mpro-mediated processing of the nsp7-10/11 polyprotein, whose mature products are cofactors of the viral replicase, identifying the order of cleavages as: 1) nsp9-10, 2) nsp8-9/nsp10-11, and 3) nsp7-8. Integrative modeling based on mass spectrometry (including hydrogen-deuterium exchange and cross-linking) and X-ray scattering yielded three-dimensional models of the nsp7-10/11 polyprotein. Our data suggest that the nsp7-10/11 structure in complex with Mpro strongly resembles the unbound polyprotein, and that both polyprotein conformation and junction accessibility determine the preference and order of cleavages. Finally, we used limited proteolysis assays to characterize the effect of a series of inhibitors/binders on Mpro processing of nsp7-11 and Mpro inhibition using a polyprotein substrate.

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“…154 Finally, Kihn et al combine HDXer and enhanced-sampling MD to study the conformational heterogeneity of PhuS, the cytoplasmic heme binding protein from Pseudomonas aeruginosa. 160 They compare the native states of apo-PhuS and holo-PhuS (i.e., heme-bound PhuS) and explain differences observed in their respective HDX-MS data. These differences contradicted the fact that crystal structures of apo-and holo-PhuS were nearly identical; they were also too large to be attributed to heme binding or conformational fluctuations alone.…”

Section: ■ Electrostatic Calculationsmentioning

confidence: 99%

“…This analysis reveals that the native state of apo- PhuS encompasses several substates, some showing significant levels of unfolding. 160 ■ OTHER COMBINATIONS OF MOLECULAR FEATURES As no single molecular feature has ever appeared to correlate well enough with HDX protection, the literature is rich with HDX prediction models combining several molecular features. The phenomenological expression proposed by Vendruscolo et al was among the first ones, 144 but many have followed.…”

Section: ■ Electrostatic Calculationsmentioning

confidence: 99%

Computational Modeling of Molecular Structures Guided by Hydrogen-Exchange Data

Devaurs

Antunes

Borysik

2022

J. Am. Soc. Mass Spectrom.

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Data produced by hydrogen-exchange monitoring experiments have been used in structural studies of molecules for several decades. Despite uncertainties about the structural determinants of hydrogen exchange itself, such data have successfully helped guide the structural modeling of challenging molecular systems, such as membrane proteins or large macromolecular complexes. As hydrogen-exchange monitoring provides information on the dynamics of molecules in solution, it can complement other experimental techniques in so-called integrative modeling approaches. However, hydrogen-exchange data have often only been used to qualitatively assess molecular structures produced by computational modeling tools. In this paper, we look beyond qualitative approaches and survey the various paradigms under which hydrogen-exchange data have been used to quantitatively guide the computational modeling of molecular structures. Although numerous prediction models have been proposed to link molecular structure and hydrogen exchange, none of them has been widely accepted by the structural biology community. Here, we present as many hydrogen-exchange prediction models as we could find in the literature, with the aim of providing the first exhaustive list of its kind. From purely structure-based models to so-called fractional-population models or knowledge-based models, the field is quite vast. We aspire for this paper to become a resource for practitioners to gain a broader perspective on the field and guide research toward the definition of better prediction models. This will eventually improve synergies between hydrogen-exchange monitoring and molecular modeling.

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Modeling the native ensemble of PhuS using enhanced sampling MD and HDX-ensemble reweighting

Cited by 12 publications

References 49 publications

Interpreting hydrogen-deuterium exchange experiments with molecular simulations: Tutorials and applications of the HDXer ensemble reweighting software [Article v1.0]

Interpreting hydrogen-deuterium exchange experiments with molecular simulations: Tutorials and applications of the HDXer ensemble reweighting software [Article v1.0]

Biochemical and structural insights into SARS-CoV-2 polyprotein processing by Mpro

Computational Modeling of Molecular Structures Guided by Hydrogen-Exchange Data

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