Tamm-Horsfall protein in humane urine: sex-dependent differences in the excretion and N-glycosylation pattern.

Tamm-Horsfall protein (THP) is a highly N-glycosylated protein from epithelial cells of the ascending limb of Henle loop. It is secreted into the urine as part of the innate immune response against uropathogenic pathogens. As women are more likely to suffer from urinary tract infections, biomedical studies were conducted to investigate sex-differences in THP excretion, as well as differences in the THP N-glycosylation pattern. A total of 238 volunteers (92 men, 146 women, 69 with hormonal contraceptives) participated in this study, providing urine samples. Women showed a clear tendency to have higher THP concentration and excretion rates than men (p < 0.16). Regular intake of hormonal contraceptives had no significant influence on urinary THP concentration compared to no regular intake. The individual N-glycosylation pattern of THP in urine samples from randomly selected individuals (10 female, 10 male) was investigated after enzymatic release and MS analysis of the oligosaccharides. Female subjects tended to have an increased proportion of oligomannose type N-glycans and non-fucosylated glycans, whereas men had an increased proportion of fucosylated complex-type glycans. The higher level of oligomannose-type glycans in THP from women might be explained by a self-defence mechanism to overcome the higher infections pressure by the female anatomical properties.

Unprotected Galactosamine as a Dynamic Key for a Cyclochiral Lock.

The discrimination of d-galactosamine (G), representative of the amino-sugar class of compounds, has been probed through nano-ESI-FT-ICR mass spectrometry by isolating the relevant [C·H·G]+ proton-bound complexes with the enantiomers of the cyclochiral resorcin[4]arene C and allowing them to react toward three primary amines (B = EtNH2, iPrNH2, and (R)- and (S)-sBuNH2). The system under investigation presents several features that help to unveil the behavior of unprotected G in such a supramolecular architecture: (i) the hydrophobic derivatization of the C convex side forces the polar guest G to be coordinated by the cyclochiral concave region; (ii) protonated d-galactosamine exists as an anomeric mixture, dynamically interconverting throughout the experimental time-window; and (iii) different basicities of B allow the experiment to subtly tune the reactivity of the [C·H·G]+ complexes. Three [C·H·G]+ aggregate-types were found to exist, differing in both their origin and reactivity. The most reactive adducts ([C·H·G]ESI+), generated in the electrospray environment, undergo a G-to-B ligand exchange in competition with a partial isomerization to the unreactive [C·H·G]GAS+-type complexes. Finally, the poorly reactive [C·H·G]SOL+ aggregates are formed in solution over an hours-long time scale. A cyclochirality effect on the reactivity was found to depend on the considered [C·H·G]+ aggregate-type.

Reactions of an anionic chelate phosphane/borata-alkene ligand with [Rh(nbd)Cl]2, [Rh(CO)2Cl]2 and [Ir(cod)Cl]2.

Borata-alkenes can serve as anionic olefin equivalent ligands in transition metal chemistry. A chelate ligand of this type is described and used for metal coordination. Deprotonation of the Mes2P(CH2)2B(C6F5)2 frustrated Lewis pair in the α-CH[B] position gave the methylene-bridged phosphane/borata-alkene anion. It reacted with the [Rh(nbd)Cl] or [Rh(CO)2Cl] dimers to give the respective neutral chelate [P/C[double bond, length as m-dash]B][Rh] complexes. The reaction of the [P/C[double bond, length as m-dash]B]- anion with [Ir(cod)Cl]2 proceeded similarly, only that the complex underwent a subsequent oxidative addition reaction at the mesityl substituent. Both the resulting Ir(iii)hydride complex 15 and the P/borata-alkene Rh system 12 were used as hydrogenation catalysts. The [P/C[double bond, length as m-dash]B(C6F5)2]Rh(nbd) complex 12 served as a catalyst for arylacetylene polymerization.

What is the role of acid-acid interactions in asymmetric phosphoric acid organocatalysis? A detailed mechanistic study using interlocked and non-interlocked catalysts.

Organocatalysis has revolutionized asymmetric synthesis. However, the supramolecular interactions of organocatalysts in solution are often neglected, although the formation of catalyst aggregates can have a strong impact on the catalytic reaction. For phosphoric acid based organocatalysts, we have now established that catalyst-catalyst interactions can be suppressed by using macrocyclic catalysts, which react predominantly in a monomeric fashion, while they can be favored by integration into a bifunctional catenane, which reacts mainly as phosphoric acid dimers. For acyclic phosphoric acids, we found a strongly concentration dependent behavior, involving both monomeric and dimeric catalytic pathways. Based on a detailed experimental analysis, DFT-calculations and direct NMR-based observation of the catalyst aggregates, we could demonstrate that intermolecular acid-acid interactions have a drastic influence on the reaction rate and stereoselectivity of asymmetric transfer-hydrogenation catalyzed by chiral phosphoric acids.

Chemical Modification and Detoxification of the Pseudomonas aeruginosa Toxin 2-Heptyl-4-hydroxyquinoline N-Oxide by Environmental and Pathogenic Bacteria.

2-Heptyl-4-hydroxyquinoline N-oxide (HQNO), a major secondary metabolite and virulence factor produced by the opportunistic pathogen Pseudomonas aeruginosa, acts as a potent inhibitor of respiratory electron transfer and thereby affects host cells as well as microorganisms. In this study, we demonstrate the previously unknown capability of environmental and pathogenic bacteria to transform and detoxify this compound. Strains of Arthrobacter and Rhodococcus spp. as well as Staphylococcus aureus introduced a hydroxyl group at C-3 of HQNO, whereas Mycobacterium abscessus, M. fortuitum, and M. smegmatis performed an O-methylation, forming 2-heptyl-1-methoxy-4-oxoquinoline as the initial metabolite. Bacillus spp. produced the glycosylated derivative 2-heptyl-1-(ß-d-glucopyranosydyl)-4-oxoquinoline. Assaying the effects of these metabolites on cellular respiration and on quinol oxidase activity of membrane fractions revealed that their EC50 values were up to 2 orders of magnitude higher than that of HQNO. Furthermore, cellular levels of reactive oxygen species were significantly lower in the presence of the metabolites than under the influence of HQNO. Therefore, the capacity to transform HQNO should lead to a competitive advantage against P. aeruginosa. Our findings contribute new insight into the metabolic diversity of bacteria and add another layer of complexity to the metabolic interactions which likely contribute to shaping polymicrobial communities comprising P. aeruginosa.

Diastereoselective anodic hetero- and homo-coupling of menthol-, 8-methylmenthol- and 8-phenylmenthol-2-alkylmalonates.

Diastereoselective radical coupling was achieved with chiral auxiliaries. The radicals were generated by anodic decarboxylation of five malonic acid derivatives. These were prepared from benzyl malonates and four menthol auxiliaries. Coelectrolyses with 3,3-dimethylbutanoic acid in methanol at platinum electrodes in an undivided cell afforded hetero-coupling products in 22-69% yield with a diastereoselectivity ranging from 5 to 65% de. Electrolyses without a coacid led to diastereomeric homo-coupling products in 21-50% yield with ratios of diastereomers being 1.17:2.00:0.81 to 7.03:2.00. The stereochemistry of the new stereogenic centers was confirmed by X-ray structure analysis and 13C NMR data.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry imaging of ochratoxin A and fumonisins in mold-infected food.

RATIONALE: Mycotoxins are toxic secondary metabolites produced by various fungi. Their distribution within contaminated material is of high interest to obtain insight into infection mechanisms and the possibility of reducing contamination during food processing. METHODS: Various vegetable foodstuffs were infected with fungi of the genera Fusarium and Aspergillus. The localization of the produced mycotoxins was studied by matrix-assisted laser desorption/ionization time-of flight mass spectrometry imaging (MALDI-MSI) of cryosections obtained from infected material. The results were confirmed by high-performance liquid chromatography/electrospray ionization triple quadrupole mass spectrometry (HPLC/MS/MS). RESULTS: The mycotoxins ochratoxin A (OTA) and fumonisins of the B- and C-series (FB1 , FB2 , FB3 , FB4 , FC1 , FC2/3 , and FC4 ) as well as partially hydrolyzed fumonisins (pHFB1 , pHFB2 , pHFB3 , pHFC1 , and pHFC2/3 ) could successfully be detected by MALDI-MSI in mold-infested foodstuffs. The toxins are distributed differently in the material: OTA is co-localized with visible fungal spoilage while fumonisins could be detected throughout the whole sample. CONCLUSIONS: This work shows the applicability of MALDI-MSI to mycotoxin analysis. It has been demonstrated that the analyzed mycotoxins are differently distributed within moldy foodstuffs. These findings show the potential of MALDI-MSI for the localization of these hazardous compounds in various plant tissues. Copyright © 2016 John Wiley & Sons, Ltd.

Unidirectional triple hydrogen rearrangement in the radical cations of electron-rich 3-aryl-1-propanols: further evidence and limitation.

The unidirectional triple-hydrogen (3H) rearrangement of the radical cations of 3-aryl-1-propanols bearing an electron-rich substitutent in the para-position was investigated for the diastereomeric 2-(4-dimethylamino)benzylcyclohexanols and 2-(4-dimethylamino)-benzylcyclopentanols and confirmed to be a highly stereospecific feature. Whereas the standard electron ionization (EI) (70 eV) mass spectra of the trans-isomers exhibit very minor (approximately 2%-3%) albeit stereospecific peaks for the relevant C8H13N*+ ions (m/z 123), the metastable ion [mass-analyzed ion kinetic energy (MIKE)] spectra show these peaks with significant relative intensity (8%-17%). The respective cis-isomers do not undergo the 3H rearrangement, be it under standard or under metastable-ion conditions. The stereospecific 3H rearrangement is suppressed in the radical cations of cis- and trans-3-(4-dimethylamino)phenylcyclohexanol, the mass and MIKE spectra of which are governed by cleavage processes of the cyclohexane ring, which impedes the stereochemical assignment of the isomers by mass spectrometry. A multistep mechanism for the unidirectional 3H rearrangement is discussed in view of the present and previous experimental data.

Enantioselective supramolecular devices in the gas phase. Resorcin[4]arene as a model system.

This review describes the state-of-art in the field of the gas-phase reactivity of diastereomeric complexes formed between a chiral artificial receptor and a biologically active molecule. The presented experimental approach is a ligand-displacement reaction carried out in a nano ESI-FT-ICR instrument, supported by a thermodynamic MS-study and molecular-mechanics and molecular-dynamics (MM/MD) computational techniques. The noncovalent ion-molecule complexes are ideal for the study of chiral recognition in the absence of complicating solvent and counterion effects.

Cyclochiral resorcin[4]arenes as effective enantioselectors in the gas phase.

The effect of cyclochirality of rccc-2,8,14,20-tetra-n-decyl-4,10,16,22-tetra-O-methylresorcin[4]arene (C) on the enantiodiscrimination of a number of chiral bidentate and tridentate aromatic and aliphatic biomolecules (G) has been investigated by nano-electrospray ionization (nano-ESI)-Fourier transform ion cyclotron resonance mass spectrometry. The experimental approach is based on the formation of diastereomeric proton-bound [C·H·G](+) complexes by nano-ESI of solutions containing an equimolar amount of quasi-enantiomers (C) together with the chiral guest (G) and the subsequent measurement of the rate of the G substitution by the attack of several achiral and chiral amines. In general, the heterochiral complexes react faster than the homochiral ones, except when G is an aminoalcoholic neurotransmitter whose complexes, beyond that, exhibit the highest enantioselectivity. The kinetic results were further supported by both collision-induced dissociation experiments on some of the relevant [C(2) ·H·G](+) three-body species and Density functional theory (DFT) calculations performed on the most selective systems.

Unprecedented gas-phase chiroselective logic gates.

The gas-phase encounters between 2-aminobutane and proton-bound chiral resorcin[4]arene/nucleoside complexes behave in the gas phase as supramolecular "chiroselective logic gates" by releasing the nucleoside depending on the resorcin[4]arene and the 2-aminobutane configurations.

Gas-phase enantioselectivity of chiral N-linked peptidoresorc[4]arene isomers toward dipeptides.

The gas-phase enantioselectivity of cone N-linked peptidoresorc[4]arenes (generally symbolized as M) toward the homologue dipeptides (generally symbolized as A) has been evaluated by measuring the kinetics of the A release from the diastereomeric [M x H x A](+) complexes induced by (R)-(-)-2-butylamine (B). In most cases investigated, the heterochiral [M x H x A](+) complexes, namely those wherein the configuration of the A guest is opposite to that of the host M pendants, react faster (up to 5 times) than the homochiral analogues, wherein guest A guest has the same configuration of the host M pendants. The kinetic results, discussed in the light of previous MS and NMR evidence, indicate that both the efficiency and the enantioselectivity of the guest exchange reaction depend essentially on the structure and the relative stability of the diastereomeric [M x H x A](+) complexes. These, in turn, depend on the functional groups and the configuration of both the guest and the host pendants. The absence of any significant effects of the B configuration indicates that, in all systems investigated, the dipeptide guest A is predominantly located outside the host chiral cavity.

Interactions of vinca alkaloid subunits with chiral amido[4]resorcinarenes: a dynamic, kinetic, and spectroscopic study.

The stereoselectivity of the reaction between (R)-(-)-2-butylamine and the diastereomeric proton-bound complexes of (+)-catharanthine (C) or (-)-vindoline (V) with some chiral amido[4]resorcinarenes has been investigated in the gas phase by ESI-FT-ICR-MS. The reaction stereoselectivity (0.56 < k(homo)/k(hetero) < 16.9) is found to depend critically on the functional groups present in the chiral pendants of the hosts. Rationalisation of the kinetic results is based on careful computational and spectroscopic studies of the most stable conformations of (+)-catharanthine and its protonated form in the isolated state and in water, as well as in a representative host structure. The emerging picture points to the relevant diastereomeric proton-bound complexes as quasi-degenerate, thus suggesting that their stereoselectivity in the guest exchange reaction is mostly due to kinetic factors. The results of this study may represent a starting point for a deeper comprehension of the intrinsic factors that endow these molecules, and their dimeric forms, with their biochemical properties.

Energetics and reaction mechanisms for the competitive losses of H2, CH4 and C2H4 from protonated methylbenzenes--implications to the methanol- to-hydrocarbons (MTH) process.

We report the unimolecular decomposition following collisional activation of protonated mono-, di- and trimethylbenzenes as a function of collision energy. The resulting energy-resolved mass spectra are then used for the quality control of high-level quantum chemical models of the respective potential energy surfaces. Distinction is made between direct dissociation products (CH(4) or H(2)) and indirect products (alkenes), since formation of the latter requires extensive rearrangement of the molecular skeleton. Very good consistency was found between model and experiment. The models thereby provide a solid foundation for discussing the reaction mechanisms of the industrial methanol-to-hydrocarbon process. The losses of CH(4), C(2)H(4) and C(3)H(6) from mesitylenium ions have been studied by (13)C and (2)H labelling and the alkene losses were found to occur via irreversible isomerisation pathways.

SIRIUS: decomposing isotope patterns for metabolite identification.

MOTIVATION: High-resolution mass spectrometry (MS) is among the most widely used technologies in metabolomics. Metabolites participate in almost all cellular processes, but most metabolites still remain uncharacterized. Determination of the sum formula is a crucial step in the identification of an unknown metabolite, as it reduces its possible structures to a hopefully manageable set. RESULTS: We present a method for determining the sum formula of a metabolite solely from its mass and the natural distribution of its isotopes. Our input is a measured isotope pattern from a high resolution mass spectrometer, and we want to find those molecules that best match this pattern. Our method is computationally efficient, and results on experimental data are very promising: for orthogonal time-of-flight mass spectrometry, we correctly identify sum formulas for >90% of the molecules, ranging in mass up to 1000 Da.

A kinetic study of guest displacement reactions on a host-guest complex with a photoswitchable calixarene.

The displacement processes of several guests, incorporated in a calixarene host system, were investigated in the gas phase by electrospray ionization-Fourier transform-ion cyclotron resonance (ESI-FT-ICR) mass spectrometry. The complexes resulting from a resorcin[4]arene host with ammonia and sec-butylamine guests were isolated in an ICR-cell, separately using both states of the photoswitch as well as two reference systems for the open and closed forms of the photoswitchable host. The isolated complexes were forced to exchange the guest by using methylamine, ethylamine and sec-butylamine, resulting in different reaction rates for all the measured systems. Especially, the reaction rates of both states of the photoswitch are dependent on the provided guest. Potential side effects like proton exchanges were examined by an H/D-exchange experiment. The results were investigated and supported by quantum chemical calculations (DFT).

Calixarenes as hosts for ammonium cations: a quantum chemical study and mass-spectrometric investigations.

Host-guest complexes of tetramethylcavitand with different ammonium cations were investigated by using a quantum chemical method at the density functional level (BP86, B3 LYP). The NH4+ cation is strongly bound to the host. Increasing methyl substitution at the cation decreases its inclination towards the complex formation. The calculated data are in line with results from electrospray ionization mass spectrometry (ESI-MS) experiments. They reveal stable aggregates only for the NH4+ cation and for the primary alkylammonium cations.

Chiral discrimination on the host-guest-complexation of resorc[4]arenes with quarternary amines.

The interaction of inherently chiral resorc[4]arenes with different chiral ammonium ions was measured by ESI-MS. For that purpose one enantiomer of the ammonium guests was labeled with deuterium to distinguish the enantiomers by their mass. We synthesized the ammonium salts by reaction of chiral primary amines with either CH3I or CD3I and analyzed the resulting ammonium iodides by NMR and optical rotation. The complexation experiments were performed by mixing the chiral host with various ratios of the unlabeled guest and its labeled enantiomer. By analysis of the integrals of the host-guest complexes we observed a chiral discrimination effect and a secondary isotope effect as well.

Inhibition of furin by serpin Spn4A from Drosophila melanogaster.

The serpin gene Spn4 from Drosophila melanogaster encodes multiple isoforms with alternative reactive site loops (RSL). Here, we show that isoform Spn4A inhibits human furin with an apparent kassoc of 5.5 x 10(6) M(-1) s(-1). The serpin forms SDS-stable complexes with the enzyme and the RSL of Spn4A is cleaved C-terminally to the sequence -Arg-Arg-Lys-Arg/ in accord with the recognition/cleavage site of furin. Immunofluorescence studies show that Spn4A is localized in the endoplasmic reticulum (ER), suggesting that the inhibitor is an interesting tool for investigating the cellular mechanisms regulating furin and for the design of agents controlling prohormone convertases.

Encapsulated guest molecules in the dimer of octahydroxypyridine[4]arene.

Recently a new type of calix[4]arenes has been synthesized via condensation of 2,6-dihydroxypyridine and a number of aldehydes. This type of pyridine[4]arenes forms capsules consisting of two single pyridine[4]arenes. These capsules can incorporate different guest molecules, like carboxylic acids and amides in this case. We proved that the guest acids really are incorporated inside the cavity of the capsules by electrospray mass spectrometry, NMR spectroscopy, and theoretical calculations.