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1.
Rapid Commun Mass Spectrom ; 33 Suppl 3: 64-71, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-29689642

RESUMO

Mass spectrometry offers an arsenal of tools for diverse proteomic investigations. This perspective article reviews some of the recent developments in the field of coupling laser-induced dissociation with mass spectrometry (LID-MS). Strategies involving labelling with a chromophore to induce specific photo-absorption properties are considered, with a focus on specific amino acid derivatization. Some of the opportunities and challenges of LID-MS after targeted labelling for increasing specificity in complex sample analysis are discussed.

2.
J Am Soc Mass Spectrom ; 29(9): 1826-1834, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29949057

RESUMO

The nonapeptide oxytocin (OT) is used as a model sulfur-containing peptide to study the damage induced by vacuum UV (VUV) radiations. In particular, the effect of the presence (or absence in reduced OT) of oxytocin's internal disulfide bridge is evaluated in terms of photo-fragmentation yield and nature of the photo-fragments. Intact, as well as reduced, OT is studied as dianions and radical anions. Radical anions are prepared and photo-fragmented in two-color experiments (UV + VUV) in a linear ion trap. VUV photo-fragmentation patterns are analyzed and compared, and radical-induced mechanisms are proposed. The effect of VUV is principally to ionize but secondary fragmentation is also observed. This secondary fragmentation seems to be considerably enabled by the initial position of the radical on the molecule. In particular, the possibility to form a radical on free cysteines seems to increase the susceptibility to VUV fragmentation. Interestingly, disulfide bridges, which are fundamental for protein structure, could also be responsible for an increased resistance to ionizing radiations. Graphical Abstract.


Assuntos
Ânions/química , Dissulfetos/química , Ocitocina/química , Espectrometria de Massas , Oxirredução , Fotólise , Raios Ultravioleta , Vácuo
3.
Anal Chem ; 90(6): 3928-3935, 2018 03 20.
Artigo em Inglês | MEDLINE | ID: mdl-29465226

RESUMO

Thanks to comprehensive and unbiased sampling of all precursor ions, the interest to move toward bottom-up proteomic with data-independent acquisition (DIA) is continuously growing. DIA offers precision and reproducibility performances comparable to true targeted methods but has the advantage of enabling retrospective data testing with the hypothetical presence of new proteins of interest. Nonetheless, the chimeric nature of DIA MS/MS spectra inherent to concomitant transmission of a multiplicity of precursor ions makes the confident identification of peptides often challenging, even with spectral library-based extraction strategy. The introduction of specificity at the fragmentation step upon ultraviolet or visible laser-induced dissociation (LID) range targeting only the subset of cysteine-containing peptides (Cys-peptide) has been proposed as an option to streamline and reduce the search space. Here, we describe the first coupling between DIA and visible LID at 473 nm to test for the presence of Cys-peptides with a peptide-centric approach. As a test run, a spectral library was built for a pool of Cys-synthetic peptides used as surrogates of human kinases (1 peptide per protein). By extracting ion chromatograms of query standard and kinase peptides spiked at different concentration levels in an Escherichia coli proteome lysate, DIA-LID demonstrates a dynamic range of detection of at least 3 decades and coefficients of precision better than 20%. Finally, the spectral library was used to search for endogenous kinases in human cellular extract.


Assuntos
Cisteína/análise , Peptídeos/química , Proteínas Quinases/química , Proteômica/métodos , Espectrometria de Massas em Tandem/métodos , Linhagem Celular , Humanos , Proteoma/química , Software , Fluxo de Trabalho , p-Dimetilaminoazobenzeno/análogos & derivados , p-Dimetilaminoazobenzeno/química
4.
J Am Soc Mass Spectrom ; 29(2): 270-283, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-28980177

RESUMO

Mass spectrometry-based methods have made significant progress in characterizing post-translational modifications in peptides and proteins; however, certain aspects regarding fragmentation methods must still be improved. A good technique is expected to provide excellent sequence information, locate PTM sites, and retain the labile PTM groups. To address these issues, we investigate 10.6 µm IRMPD, 213 nm UVPD, and combined UV and IR photodissociation, known as HiLoPD (high-low photodissociation), for phospho-, sulfo-, and glyco-peptide cations. IRMPD shows excellent backbone fragmentation and produces equal numbers of N- and C-terminal ions. The results reveal that 213 nm UVPD and HiLoPD methods can provide diverse backbone fragmentation producing a/x, b/y, and c/z ions with excellent sequence coverage, locate PTM sites, and offer reasonable retention efficiency for phospho- and glyco-peptides. Excellent sequence coverage is achieved for sulfo-peptides and the position of the SO3 group can be pinpointed; however, widespread SO3 losses are detected irrespective of the methods used herein. Based on the overall performance achieved, we believe that 213 nm UVPD and HiLoPD can serve as alternative options to collision activation and electron transfer dissociations for phospho- and glyco-proteomics. Graphical Abstract ᅟ.


Assuntos
Glicopeptídeos/química , Peptídeos/química , Fosfopeptídeos/química , Processamento de Proteína Pós-Traducional , Enxofre/análise , Sequência de Aminoácidos , Raios Infravermelhos , Espectrometria de Massas/métodos , Fotólise , Raios Ultravioleta
5.
Rapid Commun Mass Spectrom ; 31(23): 1985-1992, 2017 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-28884878

RESUMO

RATIONALE: Tandem mass spectrometry (MS/MS) is the pivotal tool for protein structural characterization and quantification. Identification relies on the fragmentation step of tryptic peptides in bottom-up strategy. Specificity of fragmentation can be obtained using laser-induced dissociation (LID) in the visible range, after tagging of the targeted peptides with an adequate chromophore. Backbone fragmentation is required to obtain specific fragments and confident identification. We present herein a study of fragmentation patterns of chromophore-tagged peptides in LID, showing the potential of LID methodology to provide the maximum number of fragments for further identification and quantification. METHODS: A total of 401 cysteine-containing tryptic peptides originating from the human proteome were derivatizated on the thiol group of cysteine with a Dabcyl maleimide chromophore, which has a high photo-absorption cross section at 473 nm. The derivatized peptides were then analyzed by LID at 473 nm on a Q Exactive instrument. RESULTS: LID spectra present a characteristic fragment at m/z 252.112 for all precursors. This product ion arises from the internal dissociation of the Dabcyl chromophore. Several peptide-backbone fragment ions are also detected. Results show the quasi absence of fragmentation at the cysteine site. This indicates that part of the energy must be redistributed across the entire system despite excitation initially localized at the chromophore. Indeed, the fragmentation mainly occurs at 3 to 5 amino acids from the derivatized cysteine residue. CONCLUSIONS: LID of derivatized cysteine-containing peptides displays the initial fragmentation of the chromophore. As energy is redistributed all along the peptide sequence, fragmentation of the peptide backbone is also observed. Thus, LID of chromophore-tagged peptides produces adequate fragment ions, allowing both good sequence coverage for a greater confidence of identification, and a large choice of transitions for specific quantification.

6.
J Am Soc Mass Spectrom ; 28(10): 2124-2131, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28744770

RESUMO

Ionization efficiency and mechanism in ESI is strongly affected by the properties of mobile phase. The use of mobile-phase properties to accurately describe droplets in ESI source is convenient but may be inadequate as the composition of the droplets is changing in the plume due to electrochemical reactions occurring in the needle tip as well as continuous drying and fission of droplets. Presently, there is paucity of research on the effect of the polarity of the ESI mode on mobile phase composition in the droplets. In this paper, the change in the organic solvent content, pH, and droplet size are studied in the ESI plume in both ESI+ and ESI- ionization mode. We introduce a rigorous way - the absolute pH (pHabsH2O) - to describe pH change in the plume that takes into account organic solvent content in the mobile phase. pHabsH2O enables comparing acidities of ESI droplets with different organic solvent contents. The results are surprisingly similar for both ionization modes, indicating that the dynamics of the change of mobile-phase properties is independent from the ESI mode used. This allows us to conclude that the evolution of ESI droplets first of all proceeds via the evaporation of the organic modifier and to a lesser extent via fission of smaller droplets from parent droplets. Secondly, our study shows that qualitative findings related to the ESI process obtained on the ESI+ mode can almost directly be applied also in the ESI- mode. Graphical Abstract ᅟ.

7.
Anal Chem ; 89(11): 5665-5668, 2017 06 06.
Artigo em Inglês | MEDLINE | ID: mdl-28489356

RESUMO

For the first time, the electrospray ionization efficiency (IE) scales in positive and negative mode are united into a single system enabling direct comparison of IE values across ionization modes. This is made possible by the use of a reference compound that ionizes to a similar extent in both positive and negative modes. Thus, choosing the optimal (i.e., most sensitive) ionization conditions for a given set of analytes is enabled. Ionization efficiencies of 33 compounds ionizing in both modes demonstrate that, contrary to general practice, negative mode allows better sensitivity for 46% of such compounds whereas the positive mode is preferred for only 18%, and for 36%, the results for both modes are comparable.

8.
J Am Soc Mass Spectrom ; 27(9): 1435-42, 2016 09.
Artigo em Inglês | MEDLINE | ID: mdl-27287047

RESUMO

Herein we report the successful implementation of the consecutive and simultaneous photodissociation with high (213 nm) and low (10.6 µm) energy photons (HiLoPD, high-low photodissociation) on ubiquitin in a quadrupole-Orbitrap mass spectrometer. Absorption of high-energy UV photon is dispersed over the whole protein and stimulates extensive C-Cα backbone fragmentation, whereas low-energy IR photon gradually increases the internal energy and thus preferentially dissociates the most labile amide (C-N) bonds. We noticed that simultaneous irradiation of UV and IR lasers on intact ubiquitin in a single MS/MS experiment provides a rich and well-balanced fragmentation array of a/x, b/y, and z ions. Moreover, secondary fragmentation from a/x and z ions leads to the formation of satellite side-chain ions (d, v, and w) and can help to distinguish isomeric residues in a protein. Implementation of high-low photodissociation in a high-resolution mass spectrometer may offer considerable benefits to promote a comprehensive portrait of protein characterization. Graphical Abstract ᅟ.

9.
J Phys Chem Lett ; 7(13): 2586-90, 2016 Jul 07.
Artigo em Inglês | MEDLINE | ID: mdl-27327376

RESUMO

Single-photon, two-electron photodetachment from nickel phthalocyanine tetrasulfonic acid tetra anions, [NiPc](4-), was examined in the gas-phase using a linear ion trap coupled to the DESIRS VUV beamline of the SOLEIL Synchrotron. This system was chosen since it has a low detachment energy, known charge localization, and well-defined geometrical and electronic structures. A threshold for two-electron loss is observed at 10.2 eV, around 1 eV lower than previously observed double detachment thresholds on multiple charged protein anions. The photodetachment energy of [NiPc](4-) has been previously determined to be 3.5 eV and the photodetachment energy of [NiPc](3-•) is determined in this work to be 4.3 eV. The observed single photon double electron detachment threshold is hence 5.9 eV higher than the energy required for sequential single electron loss. Possible mechanisms are for double photodetachment are discussed. These observations pave the way toward new, exciting experiments for probing double photodetachment at relatively low energies, including correlation measurements on emitted photoelectrons.

10.
Nat Commun ; 7: 11746, 2016 06 06.
Artigo em Inglês | MEDLINE | ID: mdl-27265868

RESUMO

Metalloenzymes preorganize the reaction environment to steer substrate(s) along the required reaction coordinate. Here, we show that phosphine ligands selectively facilitate protonation of binuclear silver hydride cations, [LAg2(H)](+) by optimizing the geometry of the active site. This is a key step in the selective, catalysed extrusion of carbon dioxide from formic acid, HO2CH, with important applications (for example, hydrogen storage). Gas-phase ion-molecule reactions, collision-induced dissociation (CID), infrared and ultraviolet action spectroscopy and computational chemistry link structure to reactivity and mechanism. [Ag2(H)](+) and [Ph3PAg2(H)](+) react with formic acid yielding Lewis adducts, while [(Ph3P)2Ag2(H)](+) is unreactive. Using bis(diphenylphosphino)methane (dppm) reshapes the geometry of the binuclear Ag2(H)(+) scaffold, triggering reactivity towards formic acid, to produce [dppmAg2(O2CH)](+) and H2. Decarboxylation of [dppmAg2(O2CH)](+) via CID regenerates [dppmAg2(H)](+). These gas-phase insights inspired variable temperature NMR studies that show CO2 and H2 production at 70 °C from solutions containing dppm, AgBF4, NaO2CH and HO2CH.


Assuntos
Dióxido de Carbono/química , Formiatos/química , Prata/química , Catálise , Descarboxilação , Íons , Ligantes , Teoria Quântica , Soluções , Espectrofotometria Infravermelho , Espectrofotometria Ultravioleta , Termodinâmica
11.
Langmuir ; 32(16): 4052-8, 2016 04 26.
Artigo em Inglês | MEDLINE | ID: mdl-27042942

RESUMO

Nanoparticle-based temperature imaging is an emerging field of advanced applications. Herein, the sensitivity of the fluorescence of rhodamine B-doped latex nanoparticles toward temperature is described. Submicrometer size latex particles were prepared by a surfactant-free emulsion polymerization method that allowed a simple and inexpensive way to incorporate rhodamine B into the nanoparticles. Also, rhodamine B-coated latex nanoparticles dispersed in water were prepared in order to address the effect of the dye location in the nanoparticles on their temperature dependence. A better linearity of the temperature dependence emission of the rhodamine B-embedded latex particles, as compared to that of free rhodamine B dyes or rhodamine B-coated latex particles, is observed. Temperature-dependent fluorescence measurements by fluorescent confocal microscopy on individual rhodamine B-embedded latex particles were found similar to those obtained for fluorescent latex nanoparticles in solution, indicating that these nanoparticles could be good candidates to probe thermal processes as nanothermometers.

12.
J Am Soc Mass Spectrom ; 27(3): 474-86, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26545767

RESUMO

Characterization of acidic peptides and proteins is greatly hindered due to lack of suitable analytical techniques. Here we present the implementation of 213 nm ultraviolet photodissociation (UVPD) in high-resolution quadrupole-Orbitrap mass spectrometer in negative polarity for peptide anions. Radical-driven backbone fragmentation provides 22 distinctive fragment ion types, achieving the complete sequence coverage for all reported peptides. Hydrogen-deficient radical anion not only promotes the cleavage of Cα-C bond but also stimulates the breaking of N-Cα and C-N bonds. Radical-directed loss of small molecules and specific side chain of amino acids are detected in these experiments. Radical containing side chain of amino acids (Tyr, Ser, Thr, and Asp) may possibly support the N-Cα backbone fragmentation. Proline comprising peptides exhibit the unusual fragment ions similar to reported earlier. Interestingly, basic amino acids such as Arg and Lys also stimulated the formation of abundant b and y ions of the related peptide anions. Loss of hydrogen atom from the charge-reduced radical anion and fragment ions are rationalized by time-dependent density functional theory (TDDFT) calculation, locating the potential energy surface (PES) of ππ* and repulsive πσ* excited states of a model amide system.


Assuntos
Peptídeos/química , Fotólise , Sequência de Aminoácidos , Íons/química , Espectrometria de Massas , Modelos Moleculares , Raios Ultravioleta
13.
J Am Soc Mass Spectrom ; 26(11): 1923-30, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26246121

RESUMO

For the first time, quantitative electrospray (ESI) ionization efficiencies (IE), expressed as logIE values, obtained on different mass-spectrometric setups (four mass analyzers and four ESI sources) are compared for 15 compounds of diverse properties. The general trends of change of IE with molecular structure are the same with all experimental setups. The obtained IE scales could be applied on different setups: there were no statistically significant changes in the order of ionization efficiency and the root mean of squared differences of the logIE values of compounds between the scales compiled on different instruments were found to be between 0.21 and 0.55 log units. The results show that orthogonal ESI source geometry gives better differentiating power and additional pneumatic assistance improves it even more. It is also shown that the ionization efficiency values are transferable between different mass-spectrometric setups by three anchoring points and a linear model. The root mean square error of logIE prediction ranged from 0.24 to 0.72 depending on the instrument. This work demonstrates for the first time the inter-instrument transferability of quantitative electrospray ionization efficiency data. Graphical Abstract ᅟ.

14.
Anal Chem ; 87(16): 8210-7, 2015 Aug 18.
Artigo em Inglês | MEDLINE | ID: mdl-26110558

RESUMO

We investigated how the temperature and size of charged droplets are affected by the electrospray ionization (ESI) process, using in situ measurements involving laser-induced fluorescence and Mie scattering on a thermal gradient focusing ESI source. Rhodamine dyes were employed as temperature indicators using ratiometric intensity-based fluorescence techniques. The results were compared to lifetime-based techniques using tris(2,2'-bipyridyl)dichlororuthenium(II) hexahydrate, [Ru(bpy)3](2+). Both methods gave similar profiles. Nevertheless, the precision and sensitivity were higher for lifetime-based techniques in comparison to intensity-based techniques. Global warming (with ΔT ∼10 K) of the ESI plume is reported while the size of the droplet decreases along the plume. The global warming indicates that the conductive thermal transfer (between the superheated sheath gas and the solvent) is predominant and stronger than the cooling effect due to the evaporation of the droplets, and this outcome is effectively reproduced by a diffusion-controlled evaporation model. Thermal gradient focusing ESI sources therefore appear to be efficient sources for evaporating large amounts of solvent, along with an increase in temperature.

15.
J Am Soc Mass Spectrom ; 26(3): 432-43, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25503080

RESUMO

UV photodissociation of proline-containing peptide ions leads to unusual product ions. In this paper, we report laser-induced dissociation of a series of proline-containing peptides at 213 nm. We observe specific fragmentation pathways corresponding to the formation of (y-2), (a + 2) and (b + 2) fragment ions. This was not observed at 266 nm or for peptides which do not contain proline residues. In order to obtain insights into the fragmentation dynamics at 213 nm, a small peptide (RPK for arginine-proline-lysine) was studied both theoretically and experimentally. Calculations of absorption spectra and non-adiabatic molecular dynamics (MD) were made. Second and third excited singlet states, S(2) and S(3), lie close to 213 nm. Non-adiabatic MD simulation starting from S(2) and S(3) shows that these transitions are followed by C-C and C-N bond activation close to the proline residue. After this first relaxation step, consecutive rearrangements and proton transfers are required to produce unusual (y-2), (a + 2) and (b + 2) fragment ions. These fragmentation mechanisms were confirmed by H/D exchange experiments.


Assuntos
Peptídeos/química , Peptídeos/efeitos da radiação , Fotólise , Prolina/química , Raios Ultravioleta , Medição da Troca de Deutério , Simulação de Dinâmica Molecular
16.
Chemistry ; 20(50): 16626-33, 2014 Dec 08.
Artigo em Inglês | MEDLINE | ID: mdl-25324009

RESUMO

Multistage mass spectrometry and density functional theory (DFT) were used to characterise the small silver hydride nanocluster, [Ag3 H2 L](+) (where L=(Ph2 P)2 CH2 ) and its gas-phase unimolecular chemistry. Collision-induced dissociation (CID) yields [Ag2 HL](+) as the major product while laser-induced dissociation (LID) proceeds via H2 formation and subsequent release from [Ag3 H2 L](+) , giving rise to [Ag3 L](+) as the major product. Deuterium labelling studies on [Ag3 D2 L](+) prove that the source of H2 is from the hydrides and not from the ligand. Comparison of TD-DFT absorption patterns obtained for the optimised structures with action spectroscopy results, allows assignment of the measured features to structures of precursors and products. Molecular dynamics "on the fly" reveal that AgH loss is favoured in the ground state, but H2 formation and loss is preferred in the first excited state S1 , in agreement with CID and LID experimental findings. This indicates favourable photo-induced formation of H2 and subsequent release from [Ag3 H2 L](+) , an important finding in context of metal hydrides as a hydrogen storage medium, which can subsequently be released by heating or irradiation with light.

17.
Analyst ; 139(21): 5523-30, 2014 Nov 07.
Artigo em Inglês | MEDLINE | ID: mdl-25197743

RESUMO

Improvement of the fragmentation specificity may streamline data processing of bottom-up proteomic experiments by drastically reducing either the amount of MS/MS data to process in the discovery phase or the detection of interfering signals in targeted quantification. Photodissociation at appropriate wavelengths is a promising alternative technique to the non-discriminating conventional activation mode by collision. Here, we describe the implementation of visible LID at 473 nm in a Q-Exactive-Orbitrap mass spectrometer for the specific detection of cysteine-containing peptides tagged with a Dabcyl group. HCD cell DC offset and irradiation time were optimized to obtain high fragmentation yield and spectra free of contaminating CID product ions, while keeping the irradiation time scale compatible with chromatographic separation. With this optimized experimental set-up, the selective detection of cysteine-containing peptides in a whole tryptic hydrolysate of three combined proteins is demonstrated by comparing all ion fragmentation (AIF) spectra recorded online with and without laser irradiation.


Assuntos
Cisteína/análise , Peptídeos/análise , Espectrometria de Massas em Tandem/métodos , Cromatografia Líquida de Alta Pressão , Peptídeos/química , Processos Fotoquímicos , Proteômica
18.
Rapid Commun Mass Spectrom ; 28(19): 2084-8, 2014 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-25156598

RESUMO

RATIONALE: To demonstrate the potential use of zebra fish (Danio rerio) as a model vertebrate organism by producing two-dimensional ion images of the whole zebra fish, and being able to distinguish particular areas of interest such as the brain, spinal cord, and stomach region using a desorption electrospray ionization (DESI) ion source coupled to a linear ion trap. METHODS: Imaging experiments are performed on 45 µm sagittal slices of zebra fish (Danio rerio), which are thaw-mounted onto microscope glass slides. The slides are then analyzed using a solvent of acetonitrile/dimethylformamide (50:50) (ACN/DMF), with a solvent flow rate of 1.5 µL/min; data are acquired in negative ion mode. Raw mass spectrum data files are converted into a readable file for Biomap. The images produced are then analyzed for ion distributions. RESULTS: We are able to create clear, distinct, chemical intensity images of the brain, spinal cord, and stomach based on lipid content as well as bile salt. The identities of these compounds were confirmed by tandem mass spectrometric (MS/MS) experiments and comparisons with literature. CONCLUSIONS: Imaging of whole zebra fish is possible using ambient ionization techniques such as DESI. Analyses are fast and reliable. For most of the compounds observed, the identification by MS/MS can be performed directly from the fish tissue sample.


Assuntos
Imagem Molecular/métodos , Espectrometria de Massas por Ionização por Electrospray/métodos , Imagem Corporal Total/métodos , Peixe-Zebra , Animais , Biomarcadores/química , Masculino
19.
J Am Soc Mass Spectrom ; 25(11): 1853-61, 2014 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25142325

RESUMO

Electrospray (ESI) ionization efficiencies (IE) of a set of 10 compounds differing by chemical nature, extent of ionization in solution (basicity), and by hydrophobicity (tetrapropylammonium and tetraethylammonium ion, triethylamine, 1-naphthylamine, N,N-dimethylaniline, diphenylphthalate, dimethylphtahalate, piperidine, pyrrolidine, pyridine) have been measured in seven mobile phases (three acetonitrile percentages 20%, 50%, and 80%, and three different pH-adjusting additives, 0.1% formic acid, 1 mM ammonia, pH 5.0 buffer combination) using the relative measurement method. MS parameters were optimized separately for each ion. The resulting relative IE data were converted into comparable logIE values by anchoring them to the logIE of tetrapropylammonium ion taking into account the differences of ionization in different solvents and thereby making the logIE values of the compounds comparable across solvents. The following conclusions were made from analysis of the data. The compounds with pK(a) values in the range of the solution pH values displayed higher IE at lower pH. The sensitivity of IE towards pH depends on hydrophobicity being very strong with pyridine, weaker with N,N-dimethylaniline, and weakest with 1-naphthylamine. IEs of tetraalkylammonium ions and triethylamine were expectedly insensitive towards solution pH. Surprisingly high IEs of phthalate esters were observed. The differences in solutions with different acetonitrile content and similar pH were smaller compared with the pH effects. These results highlight the importance of hydrophobicity in electrospray and demonstrate that high hydrophobicity can sometimes successfully compensate for low basicity.

20.
PLoS One ; 9(7): e101642, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24999730

RESUMO

Accumulation of oxidative damage in proteins correlates with aging since it can cause irreversible and progressive degeneration of almost all cellular functions. Apparently, native protein structures have evolved intrinsic resistance to oxidation since perfectly folded proteins are, by large most robust. Here we explore the structural basis of protein resistance to radiation-induced oxidation using chicken egg white lysozyme in the native and misfolded form. We study the differential resistance to oxidative damage of six different parts of native and misfolded lysozyme by a targeted tandem/mass spectrometry approach of its tryptic fragments. The decay of the amount of each lysozyme fragment with increasing radiation dose is found to be a two steps process, characterized by a double exponential evolution of their amounts: the first one can be largely attributed to oxidation of specific amino acids, while the second one corresponds to further degradation of the protein. By correlating these results to the structural parameters computed from molecular dynamics (MD) simulations, we find the protein parts with increased root-mean-square deviation (RMSD) to be more susceptible to modifications. In addition, involvement of amino acid side-chains in hydrogen bonds has a protective effect against oxidation Increased exposure to solvent of individual amino acid side chains correlates with high susceptibility to oxidative and other modifications like side chain fragmentation. Generally, while none of the structural parameters alone can account for the fate of peptides during radiation, together they provide an insight into the relationship between protein structure and susceptibility to oxidation.


Assuntos
Muramidase/química , Muramidase/metabolismo , Sequência de Aminoácidos , Raios gama , Espectrometria de Massas , Simulação de Dinâmica Molecular , Dados de Sequência Molecular , Oxirredução/efeitos da radiação , Dobramento de Proteína , Estrutura Terciária de Proteína
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