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Chemical Reviews May 2022Proton detection developed in the last 20 years as the method of choice to study biomolecules in the solid state. In perdeuterated proteins, proton dipolar interactions... (Review)
Review
Proton detection developed in the last 20 years as the method of choice to study biomolecules in the solid state. In perdeuterated proteins, proton dipolar interactions are strongly attenuated, which allows yielding of high-resolution proton spectra. Perdeuteration and backsubstitution of exchangeable protons is essential if samples are rotated with MAS rotation frequencies below 60 kHz. Protonated samples can be investigated directly without spin dilution using proton detection methods in case the MAS frequency exceeds 110 kHz. This review summarizes labeling strategies and the spectroscopic methods to perform experiments that yield assignments, quantitative information on structure, and dynamics using perdeuterated samples. Techniques for solvent suppression, H/D exchange, and deuterium spectroscopy are discussed. Finally, experimental and theoretical results that allow estimation of the sensitivity of proton detected experiments as a function of the MAS frequency and the external field in a perdeuterated environment are compiled.
Topics: Magnetic Resonance Imaging; Magnetic Resonance Spectroscopy; Nuclear Magnetic Resonance, Biomolecular; Proteins; Protons
PubMed: 34870415
DOI: 10.1021/acs.chemrev.1c00681 -
The Analyst Jun 2021Ultraviolet photodissociation (UVPD) is a powerful and rapidly developing method in top-down proteomics. Sequence coverages can exceed those obtained with collision- and...
Ultraviolet photodissociation (UVPD) is a powerful and rapidly developing method in top-down proteomics. Sequence coverages can exceed those obtained with collision- and electron-induced fragmentation methods. Because of the recent interest in UVPD, factors that influence protein fragmentation and sequence coverage are actively debated in the literature. Here, we performed top-down 213 nm UVPD experiments on a 7 T Fourier-transform ion cyclotron resonance mass spectrometer (FT-ICR MS) for the model proteins ubiquitin, myoglobin and cytochrome c that were electrosprayed from native, denaturing and supercharging solutions in order to investigate the effect of protein charge states on UVPD fragments. By performing UVPD in ultrahigh vacuum, factors associated with collisional cooling and any ion activation during transfer between mass analyzers can be largely eliminated. Sequence coverage increased from <10% for low charge states to >60% for high charge states for all three proteins. This trend is influenced by the overall charge state, i.e., charges per number of amino acid residues, and to a lesser degree by associated structural changes of protein ions of different charge states based on comparisons to published collision-cross section measurements. To rationalize this finding, and correlate sequence ion formation and identity with the number and location of protons, UVPD results were compared to protonation sites predicted based on electrostatic modelling. Assuming confined protonation sites, these results indicate the presence of two general fragmentation types; i.e., charge remote and charge directed. For moderately high protein charge states, fragment ions mostly originate in regions between likely protonation sites (charge remote), whereas sequence ions of highly charge protein ions occur either near backbone amide protonation sites at low-basicity residues (charge directed) or at charge remote sites (i.e., high-basicity residues). Overall, our results suggest that top-down 213 UVPD performance in the zero-pressure limit depends strongly on protein charge states and protonation sites can influence the location of backbone cleavages.
Topics: Ions; Mass Spectrometry; Proteomics; Protons; Ultraviolet Rays
PubMed: 34009215
DOI: 10.1039/d1an00571e -
Journal of the American Society For... Jun 2022Ionization of organic compounds with different structural and energetic properties including benzene derivatives, polycyclic aromatic hydrocarbons (PAHs), ketones, and...
Ionization of organic compounds with different structural and energetic properties including benzene derivatives, polycyclic aromatic hydrocarbons (PAHs), ketones, and polyenes was studied using a commercial atmospheric pressure corona discharge (APCI) ion source on a drift tube ion mobility-quadrupole-time-of-flight mass spectrometer (IM-QTOFMS). It was found that the studied cohort of compounds can be experimentally ionized via protonation, charge transfer, and hydride abstraction leading to formation of [M + H], [M], and [M - H] species, respectively. By experimentally monitoring the product ions and comparing the thermodynamic data for different ionization paths, it was proposed that NO is one of the main reactant ions (RIs) in the ion source used. Of particular focus in this work were theoretical and experimental studies of the effect of solvents frequently used for analytical applications with this ion source (acetonitrile, methanol, and chloroform) on the ionization mechanisms. In methanol, the studied compounds were observed to be ionized mainly via proton transfer while acetonitrile suppressed the protonation of compounds and enhanced their ionization via charge transfer and hydride abstraction. Use of chloroform as a solvent led to formation of CHCl as an alternative reactant ion (RI) to ionize the analytes via electrophilic substitution. Density functional theory (DFT) was used to study the different paths of ionization. The theoretical and experimental results showed that by using only the absolute thermodynamic data, the real ionization path cannot be determined and the energies of all competing processes such as charge transfer, protonation, and hydride abstraction need to be compared.
Topics: Acetonitriles; Atmospheric Pressure; Chloroform; Humans; Ions; Methanol; Protons; Solvents
PubMed: 35562191
DOI: 10.1021/jasms.2c00034 -
Journal of Chemical Information and... Jun 2023The interconversion between fumarate and succinate is fundamental to the energy metabolism of nearly all organisms. This redox reaction is catalyzed by a large family of...
The interconversion between fumarate and succinate is fundamental to the energy metabolism of nearly all organisms. This redox reaction is catalyzed by a large family of enzymes, fumarate reductases and succinate dehydrogenases, using hydride and proton transfers from a flavin cofactor and a conserved Arg side-chain. These flavoenzymes also have substantial biomedical and biotechnological importance. Therefore, a detailed understanding of their catalytic mechanisms is valuable. Here, calibrated electronic structure calculations in a cluster model of the active site of the Fcc fumarate reductase were employed to investigate various reaction pathways and possible intermediates in the enzymatic environment and to dissect interactions that contribute to catalysis of fumarate reduction. Carbanion, covalent adduct, carbocation, and radical intermediates were examined. Significantly lower barriers were obtained for mechanisms via carbanion intermediates, with similar activation energies for hydride and proton transfers. Interestingly, the carbanion bound to the active site is best described as an enolate. Hydride transfer is stabilized by a preorganized charge dipole in the active site and by the restriction of the C1-C2 bond in a twisted conformation of the otherwise planar fumarate dianion. But, protonation of a fumarate carboxylate and quantum tunneling effects are not critical for catalysis of the hydride transfer. Calculations also suggest that the driving force for enzyme turnover is provided by regeneration of the catalytic Arg, either coupled with flavin reduction and decomposition of a proposed transient state or directly from the solvent. The detailed mechanistic description of enzymatic reduction of fumarate provided here clarifies previous contradictory views and provides new insights into catalysis by essential flavoenzyme reductases and dehydrogenases.
Topics: Protons; Oxidation-Reduction; Catalysis; Succinates; Fumarates; Flavins; Kinetics
PubMed: 37196341
DOI: 10.1021/acs.jcim.3c00292 -
Journal of the American Chemical Society Jan 2023The catalytic transformation of N to NH by transition metal complexes is of great interest and importance but has remained a challenge to date. Despite the essential...
The catalytic transformation of N to NH by transition metal complexes is of great interest and importance but has remained a challenge to date. Despite the essential role of vanadium in biological N fixation, well-defined vanadium complexes that can catalyze the conversion of N to NH are scarce. In particular, a V(NH) intermediate derived from proton/electron transfer reactions of coordinated N remains unknown. Here, we report a dinitrogen-bridged divanadium complex bearing POCOP (2,6-(BuPO)-CH) pincer and aryloxy ligands, which can serve as a catalyst for the reduction of N to NH and NH. Low-temperature protonation and reduction of the dinitrogen complex afforded the first structurally characterized neutral metal hydrazido(2-) species ([V]═NNH), which mediated N conversion to NH, indicating that it is a plausible intermediate of the catalysis. DFT calculations showed that the vanadium hydrazido complex [V]═NNH possessed a N-H bond dissociation free energy (BDFE) of as high as 59.1 kcal/mol. The protonation of a vanadium amide complex ([V]-NH) with [PhNH][OTf] resulted in the release of NH and the formation of a vanadium triflate complex, which upon reduction under N afforded the vanadium dinitrogen complex. These transformations model the final steps of a vanadium-catalyzed N reduction cycle. Both experimental and theoretical studies suggest that the catalytic reaction may proceed via a distal pathway to liberate NH. These findings provide unprecedented insights into the mechanism of N reduction related to FeV nitrogenase.
Topics: Vanadium; Ammonia; Oxidation-Reduction; Nitrogenase; Protons; Catalysis
PubMed: 36596224
DOI: 10.1021/jacs.2c08000 -
Chemphyschem : a European Journal of... Feb 2022Anhydrous silicophosphoric acid glass with an approximate composition of H Si P O was synthesized and its thermal and proton-conducting properties were characterized....
Anhydrous silicophosphoric acid glass with an approximate composition of H Si P O was synthesized and its thermal and proton-conducting properties were characterized. Despite exhibiting a glass transition at 192 °C, the supercooled liquid could be handled as a solid up to 280 °C owing to its high viscosity. The glass and its melt exhibited proton conduction with a proton transport number of ∼1. Although covalent O-H bonds were weakened by relatively strong hydrogen bonding, the proton conductivity (4×10 S cm at 276 °C) was considerably lower than that of phosphoric acid. The high viscosity of the melt was due to the tight cross-linking of phosphate ion chains by six-fold-coordinated Si atoms. The low proton conductivity was attributed to the trapping of positively charged proton carriers around anionic SiO units (expressed as (SiO ) ) to compensate for the negative charges.
Topics: Electric Conductivity; Glass; Hydrogen Bonding; Protons
PubMed: 34862847
DOI: 10.1002/cphc.202100840 -
Faraday Discussions Jul 2022The Δp rule is commonly applied by chemists and crystal engineers as a guideline for the rational design of molecular salts and co-crystals. For multi-component... (Review)
Review
The Δp rule is commonly applied by chemists and crystal engineers as a guideline for the rational design of molecular salts and co-crystals. For multi-component crystals containing acid and base constituents, empirical evidence has shown that Δp > 4 almost always leads to salts, Δp < -1 almost always leads to co-crystals and Δp between -1 and 4 can be either. This paper reviews the theoretical background of the Δp rule and highlights the crucial role of solvation in determining the outcome of the potential proton transfer from acid to base. New data on the frequency of the occurrence of co-crystals and salts in multi-component crystal structures containing acid and base constituents show that the relationship between Δp and the frequency of salt/co-crystal formation is influenced by the composition of the crystal. For unsolvated co-crystals/salts, containing only the principal acid and base components, the point of 50% probability for salt/co-crystal formation occurs at Δp ≈ 1.4, while for hydrates of co-crystals and salts, this point is shifted to Δp ≈ -0.5. For acid-base crystals with the possibility for two proton transfers, the overall frequency of occurrence of any salt (monovalent or divalent) a co-crystal is comparable to that of the whole data set, but the point of 50% probability for observing a monovalent salt a divalent salt lies at Δp ≈ -4.5. Hence, where two proton transfers are possible, the balance is between co-crystals and divalent salts, with monovalent salts being far less common. Finally, the overall role played by the "crystal" solvation is illustrated by the fact that acid-base complexes in the intermediate region of Δp tip towards salt formation if ancillary hydrogen bonds can exist. Thus, the solvation strength of the lattice plays a key role in the stabilisation of the ions.
Topics: Hydrogen Bonding; Ions; Protons; Salts
PubMed: 35446321
DOI: 10.1039/d1fd00081k -
Biochimica Et Biophysica Acta.... Dec 2022Proteins that bind protons at cell membrane interfaces often expose to the bulk clusters of carboxylate and histidine sidechains that capture protons transiently and, in...
Proteins that bind protons at cell membrane interfaces often expose to the bulk clusters of carboxylate and histidine sidechains that capture protons transiently and, in proton transporters, deliver protons to an internal site. The protonation-coupled dynamics of bulk-exposed carboxylate clusters, also known as proton antennas, is poorly described. An essential open question is how water-mediated bridges between sidechains of the cluster respond to protonation change and facilitate transient proton storage. To address this question, here I studied the protonation-coupled dynamics at the proton-binding antenna of PsbO, a small extrinsinc subunit of the photosystem II complex, with atomistic molecular dynamics simulations and systematic graph-based analyses of dynamic protein and protein-water hydrogen-bond networks. The protonation of specific carboxylate groups is found to impact the dynamics of their local protein-water hydrogen-bond clusters. Regardless of the protonation state considered for PsbO, carboxylate pairs that can sample direct hydrogen bonding, or bridge via short hydrogen-bonded water chains, anchor to nearby basic or polar protein sidechains. As a result, carboxylic sidechains of the hypothesized antenna cluster are part of dynamic hydrogen bond networks that may rearrange rapidly when the protonation changes.
Topics: Carboxylic Acids; Histidine; Hydrogen Bonding; Photosystem II Protein Complex; Protons; Water
PubMed: 36116514
DOI: 10.1016/j.bbamem.2022.184052 -
Biomolecules Feb 2023Proton relay between interfacial water molecules allows rapid two-dimensional diffusion. An energy barrier, ΔGr‡, opposes proton-surface-to-bulk release. The...
Proton relay between interfacial water molecules allows rapid two-dimensional diffusion. An energy barrier, ΔGr‡, opposes proton-surface-to-bulk release. The ΔGr‡-regulating mechanism thus far has remained unknown. Here, we explored the effect interfacial charges have on ΔGr‡'s enthalpic and entropic constituents, ΔGH‡ and ΔGS‡, respectively. A light flash illuminating a micrometer-sized membrane patch of a free-standing planar lipid bilayer released protons from an adsorbed hydrophobic caged compound. A lipid-anchored pH-sensitive dye reported protons' arrival at a distant membrane patch. Introducing net-negative charges to the bilayer doubled ΔGH‡, while positive net charges decreased ΔGH‡. The accompanying variations in ΔGS‡ compensated for the ΔGH‡ modifications so that ΔGr‡ was nearly constant. The increase in the entropic component of the barrier is most likely due to the lower number and strength of hydrogen bonds known to be formed by positively charged residues as compared to negatively charged moieties. The resulting high ΔGr‡ ensured interfacial proton diffusion for all measured membranes. The observation indicates that the variation in membrane surface charge alone is a poor regulator of proton traffic along the membrane surface.
Topics: Protons; Lipid Bilayers; Membranes; Diffusion; Thermodynamics
PubMed: 36830721
DOI: 10.3390/biom13020352 -
International Journal of Molecular... Jun 2023Cytochrome c Oxidase (CcO), a membrane protein of the respiratory chain, pumps protons against an electrochemical gradient by using the energy of oxygen reduction to...
Cytochrome c Oxidase (CcO), a membrane protein of the respiratory chain, pumps protons against an electrochemical gradient by using the energy of oxygen reduction to water. The ("chemical") protons required for this reaction and those pumped are taken up via two distinct channels, named D-channel and K-channel, in a step-wise and highly regulated fashion. In the reductive phase of the catalytic cycle, both channels transport protons so that the pumped proton passes the D-channel before the "chemical" proton has crossed the K-channel. By performing molecular dynamics simulations of CcO in the O→E redox state (after the arrival of the first reducing electron) with various combinations of protonation states of the D- and K-channels, we analysed the effect of protonation on the two channels. In agreement with previous work, the amount of water observed in the D-channel was significantly higher when the terminal residue E286 was not (yet) protonated than when the proton arrived at this end of the D-channel and E286 was neutral. Since a sufficient number of water molecules in the channel is necessary for proton transport, this can be understood as E286 facilitating its own protonation. K-channel hydration shows an even higher dependence on the location of the excess proton in the K-channel. Also in agreement with previous work, the K-channel exhibits a very low hydration level that likely hinders proton transfer when the excess proton is located in the lower part of the K-channel, that is, on the N-side of S365. Once the proton has passed S365 (towards the reaction site, the bi-nuclear centre (BNC)), the amount of water in the K-channel provides hydrogen-bond connectivity that renders proton transfer up to Y288 at the BNC feasible. No significant direct effect of the protonation state of one channel on the hydration level, hydrogen-bond connectivity, or interactions between protein residues in the other channel could be observed, rendering proton conductivity in the two channels independent of each other. Regulation of the order of proton uptake and proton passage in the two channels such that the "chemical" proton leaves its channel last must, therefore, be achieved by other means of communication, such as the location of the reducing electron.
Topics: Electron Transport Complex IV; Protons; Electron Transport; Oxidation-Reduction; Water; Rhodobacter sphaeroides
PubMed: 37445646
DOI: 10.3390/ijms241310464