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Native mass spectrometry (nMS) provides insights into the structures and dynamics of biomacromolecules in their native-like states by preserving noncovalent interactions through "soft" electrospray ionization (ESI). For native proteins, the number of charges that are acquired scales with the surface area and mass. Here, we explore the effect of highly negatively charged DNA on the ESI charge of protein complexes and find a reduction of the mass-to-charge ratio as well as a greater variation. The charge state distributions of pure DNA assemblies show a lower mass-to-charge ratio than proteins due to their greater density in the gas phase, whereas the charge of protein–DNA complexes can additionally be influenced by the distribution of the ESI charges, ion pairing events, and collapse of the DNA components. Our findings suggest that structural features of protein–DNA complexes can result in lower charge states than expected for proteins. [ABSTRACT FROM AUTHOR]

We have investigated the effect of the target atomic number Z2 on the mean charge state (q¯ using various model predictions such as the Shima–Ishihara –Mikumo, Ziegler–Biersack–Littmark, Schiwietz, Schiwietz–Grande , Fermi-gas-models and theoretical codes with experimental data available in the literature. This investigation makes it possible to determine the best-fit model to calculate q¯. In this work, we discuss the post-collision charge state distribution in different targets used as thin films and projectile beams (Fq+, Siq+, Clq+, and Cuq+) with different charge states (available in literature). A detailed overview of such collision experiments has been explored over a wide energy range of 1.07–3.93 MeV/u. In this contribution, an overview of the mean charge state dependence on the Fermi velocity of target materials is provided. Finally, the influence of the non-radiative electron capture at the target exit surface on the projectile charge state distribution for fast projectiles in different targets is shown, and a comparison is made with experimental data.

We report the effect of N2 gas‐mixing in the xenon electron cyclotron resonance (ECR) plasma, and abundance‐dependent novel, exciting and unusual trends of the isotope anomaly. The xenon plasma was produced using a 10 GHz all‐permanent‐magnet NANOGAN ECR ion source, and the charge state distributions of naturally abundant six stable xenon isotopes with and without N2 gas‐mixing (at 25%, 50%, and 75%) were recorded. The intensity ratio of the heavier to lighter isotope, where the heavier isotope is less abundant, showed a clear signature of the isotope anomaly as explained by the linear Landau wave damping theory. Contrary to the theoretical prediction that the isotope anomaly should vanish with a relatively large fraction of the heavier isotope in mixed plasmas, the trends of intensity ratios observed in such cases are very unusual and have almost the mirror‐symmetrical shapes of those trends recorded with less abundant heavier isotope. Further, the effect of relative mass difference on the isotope anomaly was also evidenced. The N2 gas‐mixing of the xenon plasma at 25% and 50% shifted the entire charge state distribution toward the higher intensity side owing to the supply of additional electrons that caused high ionization efficiency. However, a prominent gas‐mixing effect was observed at 75% of N2 mixing in the xenon plasma beyond the +7 charge state. The abundance‐dependent unusual trends in isotope anomaly have been explained by considering different ionic temperatures, ion heating by the wave damping, and Coulomb scattering in the core of the plasma. [ABSTRACT FROM AUTHOR]

Liquid chromatography (LC) coupled with tandem mass spectrometry (MS/MS) provides a simple and efficient means for the measurement of analytes in biological matrices with high selectivity and specificity. LC–MS/MS plays an important role in the pharmaceutical industry and biomedical research, but it requires analytes to be in an ionized form in order to be detected. This can pose a challenge for large molecules such as proteins and peptides, because they can exist in multiple charged forms, and this will reduce the total analyte signal by distributing it into multiple ion peaks with a different number of charges in a mass spectrum. In conventional LC–MS/MS analysis of such macromolecules, one charged form is selected as the precursor ion which is then fragmented by collision-induced dissociation (CID) in MS/MS to generate product ions, a process referred to as multiple-reaction monitoring (MRM). The MRM method minimizes interference from endogenous molecules within biological matrices that share the same molecular weight of the precursor ion, but at the expense of signal intensity as compared to precursor ion intensity. We describe here an approach to boost detection sensitivity and expand dynamic range in the quantitation of large molecules while maintaining analytical specificity using summation of MRM (SMRM) transitions and LC separation technique. Protein image from PDB-101 (PDB101.rscb.org) [ABSTRACT FROM AUTHOR]

The xcalib toolkit has been developed to calibrate the beam profile of an X‐ray free‐electron laser (XFEL) at the focal spot based on the experimental charge state distributions (CSDs) of light atoms. Characterization of the fluence distribution at the focal spot is essential to perform the volume integrations of physical quantities for a quantitative comparison between theoretical and experimental results, especially for fluence‐dependent quantities. The use of the CSDs of light atoms is advantageous because CSDs directly reflect experimental conditions at the focal spot, and the properties of light atoms have been well established in both theory and experiment. Theoretical CSDs are obtained using xatom, a toolkit to calculate atomic electronic structure and to simulate ionization dynamics of atoms exposed to intense XFEL pulses, which involves highly excited multiple core‐hole states. Employing a simple function with a few parameters, the spatial profile of an XFEL beam is determined by minimizing the difference between theoretical and experimental results. The optimization procedure employing the reinforcement learning technique can automatize and organize calibration procedures which, before, had been performed manually. xcalib has high flexibility, simultaneously combining different optimization methods, sets of charge states, and a wide range of parameter space. Hence, in combination with xatom, xcalib serves as a comprehensive tool to calibrate the fluence profile of a tightly focused XFEL beam in the interaction region. [ABSTRACT FROM AUTHOR]

Abstract Pure and different support gas-mixed krypton plasmas are produced using the 14.5 GHz all permanent-magnet based electron cyclotron resonance (ECR) ion source. The ions are extracted and the charge state distributions (CSDs) are studied. The H 2 , He, N 2 , O 2 , Ne, Ar, CH 4 and CO 2 gases are used as support gases. One of the support gases are mixed with Kr at several mixing ratios and its effect on the outputs currents of the Kr ions are studied. The total effective charge, as well as effective charges of Kr and mixing gas ions are calculated from the measured ion currents. The total ion loss rates are also calculated. The gases N 2 , O 2 , Ar and CO 2 are found to exhibit better mixing effect in the enhancement of output of highly charged ions. The best enhancement in the output currents of Kr ions is observed in case of O 2 as the mixing gas under certain conditions. To the best of our knowledge such a detail comparative study on the influence of mixing gas has not been reported so far for ECR ion-beam application. [ABSTRACT FROM AUTHOR]

Absolute cross sections measured using electromagnetic devices to separate and detect heavy recoiling ions need to be corrected for charge state fractions. Accurate prediction of charge state distributions using theoretical models is not always a possibility, especially in energy and mass regions where data is sparse. As such, it is often necessary to measure charge state fractions directly. In this paper we present a novel method of using a scintillation screen along with a CMOS camera to image the charge dispersed beam after a set of magnetic dipoles. A measurement of the charge state distribution for 88Sr passing through a natural carbon foil is performed. Using a Bayesian model to extract statistically meaningful uncertainties from these images, we find agreement between the new method and a more traditional method using Faraday cups. Future work is need to better understand systematic uncertainties. Our technique offers a viable method to measure charge state distributions. [ABSTRACT FROM AUTHOR]

Abstract The charge state distributions of ions passing through gases or foils have a significant influence on the performance of AMS facilities. Most important is the stripping process in the terminal of the accelerator. Published data on charge state distributions have been collected as well as associated charge-changing cross sections for various projectiles and targets relevant to AMS. Based on these data, the stripping yields as function of the stripper thickness and ion energy are calculated. This allows to evaluate the stripping thickness required for reaching charge state equilibrium, as well as mass fractionation effects depending on energy and stripper thickness. The significantly different behavior of helium compared to other gases leads to very high stripping yields for specific charge states at low energies. The cross sections for electron detachment of negative ions give a tool for estimating beam losses in the low energy acceleration tube. Further, electron capture and loss cross sections allow the estimation of backgrounds caused by charge exchange processes in the acceleration tubes. The effects discussed here are illustrated with calculations and simulations for Carbon beams. The data are useful for the design of new AMS facilities or improving existing ones. [ABSTRACT FROM AUTHOR]

Background: Targeted quantification of protein biomarkers with liquid chromatography-tandem mass spectrometry (LC-MS/MS) has great potential, but is still in its infancy. Therefore, we elucidated the influence of charge state distribution and matrix effects on accurate quantification, illustrated by the peptide hormone hepcidin. Methods: An LC-MS/MS assay for hepcidin, developed based on existing literature, was improved by using 5 mM ammonium formate buffer as mobile phase A and as an elution solution for solid phase extraction (SPE) to optimize the charge state distribution. After extensive analytical validation, focusing on interference and matrix effects, the clinical consequence of this method adjustment was studied by performing receiving operating characteristic (ROC)-curve analysis in patients with iron deficiency anemia (IDA, n = 44), anemia of chronic disease (ACD, n = 42) and non-anemic patients (n = 93). Results: By using a buffered solution during sample preparation and chromatography, the most abundant charge state was shifted from 4+ to 3+ and the charge state distribution was strongly stabilized. The matrix effects which occurred in the 4+ state were therefore avoided, eliminating bias in the low concentration range of hepcidin. Consequently, sensitivity, specificity and positive predictive value (PPV) for detection of IDA patients with the optimized assay (96%, 97%, 91%, respectively) were much better than for the original assay (73%, 70%, 44%, respectively). Conclusions: Fundamental improvements in LC-MS/MS assays greatly impact the accuracy of protein quantification. This is urgently required for improved diagnostic accuracy and clinical value, as illustrated by the validation of our hepcidin assay. [ABSTRACT FROM AUTHOR]

Laser electrospray mass spectrometry (LEMS) measurement of the dissociation constant (Kd) for hen egg white lysozyme (HEWL) and N,N′,N″-triacetylchitotriose (NAG3) revealed an apparent Kd value of 313.2 ± 25.9 μM for the ligand titration method. Similar measurements for N,N′,N″,N″’-tetraacetylchitotetraose (NAG4) revealed an apparent Kd of 249.3 ± 13.6 μM. An electrospray ionization mass spectrometry (ESI-MS) experiment determined a Kd value of 9.8 ± 0.6 μM. In a second LEMS approach, a calibrated measurement was used to determine a Kd value of 6.8 ± 1.5 μM for NAG3. The capture efficiency of LEMS was measured to be 3.6 ± 1.8% and is defined as the fraction of LEMS sample detected after merging with the ESI plume. When the dilution is factored into the ligand titration measurement, the adjusted Kd value was 11.3 μM for NAG3 and 9.0 μM for NAG4. The calibration method for measuring Kd developed in this study can be applied to solutions containing unknown analyte concentrations.Graphical Abstract [ABSTRACT FROM AUTHOR]

The detection of lysozyme, or a mixture of lysozyme, cytochrome c, and myoglobin, from solutions with varying salt concentrations (0.1 to 250 mM NaCl) is compared using laser electrospray mass spectrometry (LEMS) and electrospray ionization-mass spectrometry (ESI-MS). Protonated protein peaks were observed up to a concentration of 250 mM NaCl in the case of LEMS. In the case of ESI-MS, a protein solution with salt concentration > 0.5 mM resulted in predominantly salt-adducted features, with suppression of the protonated protein ions. The constituents in the mixture of proteins were assignable up to 250 mM NaCl for LEMS and were not assignable above a NaCl concentration of 0.5 mM for ESI. The average sodium adducts (< n >) bound to the 7+ charge state of lysozyme for LEMS measurements from salt concentrations of 2.5, 25, 50, and 100 mM NaCl are 1.71, 5.23, 5.26, and 5.11, respectively. The conventional electrospray measurements for lysozyme solution containing salt concentrations of 0.1, 1, 2, and 5 mM NaCl resulted in < n > of 2.65, 6.44, 7.57, and 8.48, respectively. LEMS displays an approximately two orders of magnitude higher salt tolerance in comparison with conventional ESI-MS. The non-equilibrium partitioning of proteins on the surface of the charged droplets is proposed as the mechanism for the high salt tolerance phenomena observed in the LEMS measurements.ᅟ [ABSTRACT FROM AUTHOR]

Theta nanoelectrospray ionization of protein ions formed from aqueous buffer solutions that are mixed with denaturing solutions containing cyclic alkylcarbonates ( e.g. , vinyl ethylene carbonate; VEC) results in a significant increase in the extent of ion charging compared to native mass spectrometry. For six proteins, the extent of ion charging can be significantly higher than that obtained using denaturing solutions and alternative native “supercharging” methods. In theta nanoelectrospray supercharging, the extent of charging scales with protein mass in agreement with an analytical scaling relationship for ions with elongated structures. Theta nanoelectrospray supercharging of non-covalent complexes from native solutions results in essentially the complete loss of protein-ligand and protein-protein interactions. Based on circular dichroism spectroscopy, VEC can effectively denature proteins in buffered solutions. These data provide evidence that enrichment of VEC in theta nanoelectrospray ionization generated droplets can denature proteins on the timescale of droplet desolvation and ion formation. This approach can be used to form highly charged protein ions from native solutions containing biological buffers, including some that are considered incompatible with native MS. Forming some protein ions in the highest reported charge states directly from native solutions is no longer a challenge in obtaining primary structural information using tandem mass spectrometry. [ABSTRACT FROM AUTHOR]

Electrospray ionization mass spectra of biomolecules typically consist of a series of multiply charged ions because of the transfer of protons or other charge carriers between ions of biomolecules and the surrounding liquid or gas. The distribution of intensities of ions retained charge carriers contains information about the spatial structure of biomolecules. A new method is developed for the separation and decomposition of multidimensional charge distributions of ions bearing other charge carriers, such as alkali metal ions, along with protons. The proposed method ensures the estimation of the probability of charge carrier retention by separate functional groups for the selected conformations of biomolecules. The paper describes the application of this method to the analysis of a two-dimensional charge distribution of horse heart cytochrome C, resulting in the revelation of at least two its structural forms under the studied conditions. [ABSTRACT FROM AUTHOR]

Charge state distributions are measured using mass spectrometry for both native and denatured cytochrome c and myoglobin after laser vaporization from the solution state into an electrospray (ES) plume consisting of a series of solution additives differing in gas-phase basicity. The charge distribution depends on both the pH of the protein solution prior to laser vaporization and the gas-phase basicity of the solution additive employed in the ES solvent. Cytochrome c (myoglobin) prepared in solutions with pH of 7.0, 2.6, and 2.3 resulted in the average charge state distribution (Z) of 7.0 ± 0.1 (8.2 ± 0.1), 9.7 ± 0.2 (14.5 ± 0.3), and 11.6 ± 0.3 (16.4 ± 0.1), respectively, in ammonium formate ES solvent. The charge distribution shifted from higher charge states to lower charge states when the ES solvent contained amines additives with higher gas-phase basicity. In the case of triethyl ammonium formate, Z of cytochrome c (myoglobin) prepared in solutions with pH of 7.0, 2.6, and 2.3 decreased to 4.9 (5.7), 7.4 ± 0.2 (9.6 ± 0.3), and 7.9 ± 0.3 (9.8 ± 0.2), respectively. The detection of a charge state distribution corresponding to folded protein after laser vaporized, acid-denatured protein interacts with the ES solvent containing ammonium formate, ammonium acetate, triethyl ammonium formate, and triethyl ammonium acetate suggests that at least a part of protein population folds within the electrospray droplet on a millisecond timescale. [ABSTRACT FROM AUTHOR]

Lasso peptides are characterized by a mechanically interlocked structure, where the C-terminal tail of the peptide is threaded and trapped within an N-terminal macrolactam ring. Their compact and stable structures have a significant impact on their biological and physical properties and make them highly interesting for drug development. Ion mobility - mass spectrometry (IM-MS) has shown to be effective to discriminate the lasso topology from their corresponding branched-cyclic topoisomers in which the C-terminal tail is unthreaded. In fact, previous comparison of the IM-MS data of the two topologies has yielded three trends that allow differentiation of the lasso fold from the branched-cyclic structure: (1) the low abundance of highly charged ions, (2) the low change in collision cross sections (CCS) with increasing charge state and (3) a narrow ion mobility peak width. In this study, a three-dimensional plot was generated using three indicators based on these three trends: (1) mean charge divided by mass (ζ), (2) relative range of CCS covered by all protonated molecules (ΔΩ/Ω) and (3) mean ion mobility peak width (δΩ). The data were first collected on a set of twenty one lasso peptides and eight branched-cyclic peptides. The indicators were obtained also for eight variants of the well-known lasso peptide MccJ25 obtained by site-directed mutagenesis and further extended to five linear peptides, two macrocyclic peptides and one disulfide constrained peptide. In all cases, a clear clustering was observed between constrained and unconstrained structures, thus providing a new strategy to discriminate mechanically interlocked topologies. [ABSTRACT FROM AUTHOR]

We report the charge state distributions of the pure, 25% and 50% oxygen mixed krypton plasma to shed more light on the understanding of the gas mixing and the isotope anomaly [A. G. Drentje, Rev. Sci. Instrum. 63 (1992) 2875 and Y Kawai, D Meyer, A Nadzeyka, U Wolters and K Wiesemann, Plasma Sources Sci. Technol. 10 (2001) 451] in the electron cyclotron resonance (ECR) plasmas. The krypton plasma was produced using a 10 GHz all-permanent-magnet ECR ion source. The intensities of the highly abundant four isotopes, viz. 82Kr (~11.58%), 83Kr (~11.49%), 84Kr (~57%) and 86Kr (17.3%) up to ~ +14 charge state have been measured by extracting the ions from the plasma and analysing them in the mass and the energy using a large acceptance analyzer-cum-switching dipole magnet. The influence of the oxygen gas mixing on the isotopic krypton ion intensities is clearly evidenced beyond +9 charge state. With and without oxygen mixing, the charge state distribution of the krypton ECR plasma shows the isotope anomaly with unusual trends. The anomaly in the intensities of the isotopes having quite closer natural abundance, viz. 82Kr, 86Kr and 83Kr, 86Kr is prominent, whereas the intensity ratio of 86Kr to 84Kr shows a weak signature of it. The isotope anomaly tends to disappear with increasing oxygen mixing in the plasma. The observed trends in the intensities of the krypton isotopes do not follow the prediction of linear Landau wave damping in the plasma. [ABSTRACT FROM AUTHOR]