Molecular Dynamics Simulations Provide Atomistic Insight into Hydrogen Exchange Mass Spectrometry Experiments

Petruk, Ariel Alcides; Defelipe, Lucas A.; Limardo, Ramiro Gonzalo Rodriguez; Bucci, Hernán Andrés; Marti, Marcelo Adrian; Turjanski, Adrián

doi:10.1021/ct300519v

Cited by 32 publications

(43 citation statements)

References 37 publications

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“…The longest atomistic MD trajectory previously used for HX analysis was three orders of magnitude shorter (90 ns) (36). Rather than attempting to identify the O state, that study correlated the trajectory-averaged (essentially C-state) amide solvent access with the number of exchanged hydrogens (detected by mass spectrometry) for a set of peptide fragments (36). However, the strong linear correlation thus obtained seems to be spurious, largely resulting from the trivial increase of both variables with peptide size.…”

Section: Discussionmentioning

confidence: 99%

“…However, the physical significance of the PF is obscured by our ignorance about the structure and dynamics of the O state. Several attempts have been made to correlate experimental PFs with physical attributes of the amides, such as solvent contact (33)(34)(35)(36)(37), burial depth (38), intramolecular H-bonds (35,(38)(39)(40), packing density (38,41), or electric field (42). Where significant correlations have been found, they suggest that the chosen attribute can serve as a proxy for the propensity for C → O fluctuations.…”

Section: Significancementioning

confidence: 99%

See 1 more Smart Citation

How amide hydrogens exchange in native proteins

Persson

Halle

2015

Proc. Natl. Acad. Sci. U.S.A.

127

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Amide hydrogen exchange (HX) is widely used in protein biophysics even though our ignorance about the HX mechanism makes data interpretation imprecise. Notably, the open exchange-competent conformational state has not been identified. Based on analysis of an ultralong molecular dynamics trajectory of the protein BPTI, we propose that the open (O) states for amides that exchange by subglobal fluctuations are locally distorted conformations with two water molecules directly coordinated to the N-H group. The HX protection factors computed from the relative O-state populations agree well with experiment. The O states of different amides show little or no temporal correlation, even if adjacent residues unfold cooperatively. The mean residence time of the O state is ∼100 ps for all examined amides, so the large variation in measured HX rate must be attributed to the opening frequency. A few amides gain solvent access via tunnels or pores penetrated by water chains including native internal water molecules, but most amides access solvent by more local structural distortions. In either case, we argue that an overcoordinated N-H group is necessary for efficient proton transfer by Grotthuss-type structural diffusion.protein dynamics | hydration | proton transfer | MD simulation | BPTI B efore the tightly packed and densely H-bonded structure of globular proteins had been established, Hvidt and Linderstrøm-Lang (1) showed that all backbone amide hydrogens of insulin exchange with water hydrogens, implying that all parts of the polypeptide backbone are, at least transiently, exposed to solvent. In the following 60 y, hydrogen exchange (HX), usually monitored by NMR spectroscopy (2) or mass spectrometry (3), has been widely used to study protein folding and stability (4-10), structure (11, 12), flexibility and dynamics (13-15), and solvent accessibility and binding (16,17), often with single-residue resolution. However, because the exchange mechanism is unclear, HX data from proteins can, at best, be interpreted qualitatively (18)(19)(20)(21)(22)(23)(24)(25).Under most conditions, amide HX is catalyzed by hydroxide ions (26, 27) at a rate that is influenced by inductive and steric effects from adjacent side chains (28). For unstructured peptides, HX is a slow process simply because the hydroxide concentration is low. For example, at 25°C and pH 4, HX occurs on a time scale of minutes. Under similar conditions, amides buried in globular proteins exchange on a wide range of time scales, extending up to centuries. HX can only occur if the amide is exposed to solvent, so conformational fluctuations must be an integral part of the HX mechanism (18).Under sufficiently destabilizing conditions HX occurs from the denatured-state ensemble, but under native conditions few amides exchange by such global unfolding (9, 29-31). For example, in bovine pancreatic trypsin inhibitor (BPTI), 8 amides in the core β-sheet exchange by global unfolding under native conditions (7, 32), whereas the remaining 45 amides require less extensive conforma...

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Section: Discussionmentioning

confidence: 99%

Section: Significancementioning

confidence: 99%

How amide hydrogens exchange in native proteins

Persson

Halle

2015

Proc. Natl. Acad. Sci. U.S.A.

127

View full text Add to dashboard Cite

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“…Recently, Petruk et al 17 used MD simulations and a metric based on average SASA of NH and the number of H-bonds with water molecule as the basis for the binary decision if exchange would occur or not in the ERK2 MAP kinase system. The authors were successful in explaining some of the observed differences between apo and holo dynamics in terms of their metrics.…”

Section: Introductionmentioning

confidence: 99%

Estimation of Hydrogen-Exchange Protection Factors from MD Simulation Based on Amide Hydrogen Bonding Analysis

Park

Venable

Steckler

et al. 2015

J. Chem. Inf. Model.

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Hydrogen exchange (HX) studies have provided critical insight into our understanding of protein folding, structure and dynamics. More recently, Hydrogen Exchange Mass Spectrometry (HX-MS) has become a widely applicable tool for HX studies. The interpretation of the wealth of data generated by HX-MS experiments as well as other HX methods would greatly benefit from the availability of exchange predictions derived from structures or models for comparison with experiment. Most reported computational HX modeling studies have employed solvent-accessible-surface-area based metrics in attempts to interpret HX data on the basis of structures or models. In this study, a computational HX-MS prediction method based on classification of the amide hydrogen bonding modes mimicking the local unfolding model is demonstrated. Analysis of the NH bonding configurations from Molecular Dynamics (MD) simulation snapshots is used to determine partitioning over bonded and non-bonded NH states and is directly mapped into a protection factor (PF) using a logistics growth function. Predicted PFs are then used for calculating deuteration values of peptides and compared with experimental data. Hydrogen exchange MS data for Fatty acid synthase thioesterase (FAS-TE) collected for a range of pHs and temperatures was used for detailed evaluation of the approach. High correlation between prediction and experiment for observable fragment peptides is observed in the FAS-TE and additional benchmarking systems that included various apo/holo proteins for which literature data were available. In addition, it is shown that HX modeling can improve experimental resolution through decomposition of in-exchange curves into rate classes, which correlate with prediction from MD. Successful rate class decompositions provide further evidence that the presented approach captures the underlying physical processes correctly at the single residue level. This assessment is further strengthened in a comparison of residue resolved protection factor predictions for staphylococcal nuclease with NMR data, which was also used to compare prediction performance with other algorithms described in the literature. The demonstrated transferable and scalable MD based HX prediction approach adds significantly to the available tools for HX-MS data interpretation based on available structures and models.

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“…The most pronounced decrease in backbone H/D exchange was observed for peptides 3 (16-26) and 4 (23-28), both containing Arg24, and peptides 7 (37-47) and 8 (40)(41)(42)(43)(44)(45)(46)(47), both containing Arg43 from a neighboring monomer subunit (Figure 2a, c). Other regions located close to the active site [peptides 10 (50-72), 13 (95-103), and 19 (178-182)], and peptides 1 (2-10) and 10 (50-72) from adjacent subunit are under the influence of the P i .…”

Section: Phosphate Binding Is Affected By Arg24ala Mutation and Activmentioning

confidence: 97%

“…Different calculation strategies are employed to link protein dynamics with amide H/DX reaction rates by analyzing ensemble representation of the protein structure [35][36][37][38][39]. All-atom molecular dynamics (MD) simulations can be used to probe protein structure flexibility caused by frequent atomic movements [40,41] but other fluctuations relevant for H/DX occur on a time scale longer than it is currently possible to simulate with average computation resources. Recently, McAllister and Konermann carried out detailed structureexchange rate relationship analysis by combining MD simulations and H/DX data and showed that significant fraction of amide hydrogens exist with H/DX protection different than can be predicted by any of the current models proposed to explain particular H/DX pathway [42].…”

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confidence: 99%

New Insights into Active Site Conformation Dynamics of E. coli PNP Revealed by Combined H/D Exchange Approach and Molecular Dynamics Simulations

Kazazić

Bertoša

Luić

et al. 2015

J. Am. Soc. Mass Spectrom.

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Abstract. The biologically active form of purine nucleoside phosphorylase (PNP) from Escherichia coli (EC 2.4.2.1) is a homohexamer unit, assembled as a trimer of dimers. Upon binding of phosphate, neighboring monomers adopt different active site conformations, described as open and closed. To get insight into the functions of the two distinctive active site conformations, virtually inactive Arg24Ala mutant is complexed with phosphate; all active sites are found to be in the open conformation. To understand how the sites of neighboring monomers communicate with each other, we have combined H/D exchange (H/DX) experiments with molecular dynamics (MD) simulations. Both methods point to the mobility of the enzyme, associated with a few flexible regions situated at the surface and within the dimer interface. Although H/ DX provides an average extent of deuterium uptake for all six hexamer active sites, it was able to indicate the dynamic mechanism of cross-talk between monomers, allostery. Using this technique, it was found that phosphate binding to the wild type (WT) causes arrest of the molecular motion in backbone fragments that are flexible in a ligand-free state. This was not the case for the Arg24Ala mutant. Upon nucleoside substrate/inhibitor binding, some release of the phosphate-induced arrest is observed for the WT, whereas the opposite effects occur for the Arg24Ala mutant. MD simulations confirmed that phosphate is bound tightly in the closed active sites of the WT; conversely, in the open conformation of the active site of the WT phosphate is bound loosely moving towards the exit of the active site. In Arg24Ala mutant binary complex P i is bound loosely, too.

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Molecular Dynamics Simulations Provide Atomistic Insight into Hydrogen Exchange Mass Spectrometry Experiments

Cited by 32 publications

References 37 publications

How amide hydrogens exchange in native proteins

How amide hydrogens exchange in native proteins

Estimation of Hydrogen-Exchange Protection Factors from MD Simulation Based on Amide Hydrogen Bonding Analysis

New Insights into Active Site Conformation Dynamics of E. coli PNP Revealed by Combined H/D Exchange Approach and Molecular Dynamics Simulations

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