Electrospray ionization quadrupole time-of-flight (ESI-QqToF) mass spectra of the zwitteronic sal... more Electrospray ionization quadrupole time-of-flight (ESI-QqToF) mass spectra of the zwitteronic salts naloxonazine dihydrochloride 1 and naloxone hydrochloride 2, a common series of morphine opiate receptor antagonists, were recorded using different declustering potentials. The singly charged ion [M+H-2HCl](+) at m/z 651.3170 and the doubly charged ion [M+2H-2HCl](2+) at m/z 326.1700 were noted for naloxonazine dihydrochloride 1; and the singly charged ion [M+H-HCl](+) at m/z 328.1541 was observed for naloxone hydrochloride 2. Low-energy collision-induced dissociation tandem mass spectrometry (CID-MS/MS) experiments established the fragmentation routes of these compounds. In addition to the characteristic diagnostic product ions obtained, we noticed the formation of a series of radical product ions for the zwitteronic compounds 1 and 2, and also the formation of a distonic ion product formed from the singly charged ion [M+H-HCl](+) of naloxone hydrochloride 2. Confirmation of the various established fragmentation routes was effected by conducting a series of ESI-CID-QqTof-MS/MS product ion scans, which were initiated by CID in the atmospheric pressure/vacuum interface using a higher declustering potential. Deuterium labeling was also performed on the zwitteronic salts 1 and 2, in which the hydrogen atoms of the OH and NH groups were exchanged with deuterium atoms. Low-energy CID-QqTof-MS/MS product ion scans of the singly charged and doubly charged deuteriated molecules confirmed the initial fragmentation patterns proposed for the protonated molecules. Precursor ion scan analyses were also performed with a conventional quadrupole-hexapole-quadrupole tandem mass spectrometer and allowed the confirmation of the genesis of some diagnostic ions.
Recently, we identified a novel disulfide oxidoreductase, SdbA, in the oral bacterium Streptococc... more Recently, we identified a novel disulfide oxidoreductase, SdbA, in the oral bacterium Streptococcus gordonii. Disulfide oxidoreductases form disulfide bonds in nascent proteins using a CXXC catalytic motif. Typically, the N-terminal cysteine interacts with substrates, while the C-terminal cysteine is buried and only reacts with the first cysteine of the motif. In this study, we investigated the SdbA C(86) P(87) D(88) C(89) catalytic motif. In vitro, SdbA single cysteine variants at the N or C-terminal position (SdbAC86P and SdbAC89A ) were active, but displayed different susceptibility to oxidation, and N-terminal cysteine was prone to sulfenylation. In S. gordonii, mutants with a single N-terminal were inactive and formed unstable disulfide adducts with other proteins. Activity was partially restored by inactivation of pyruvate oxidase, a hydrogen peroxide generator. Presence of the C-terminal cysteine alone (in the SdbAC86P variant) could complement the ΔsdbA mutant and restore di...
NATO Science for Peace and Security Series A: Chemistry and Biology, 2010
... to 17b-estradiol in the granulose cells by the action of cytochrome P450 aromatase ... mRNA c... more ... to 17b-estradiol in the granulose cells by the action of cytochrome P450 aromatase ... mRNA can be qualitatively assayed by direct northern blotting while more sophis-ticated methods ... Vitellogenin, the egg yolk precursor protein, has become a popular biomarker for measuring ...
European journal of mass spectrometry (Chichester, England), 2003
Vitellogenin (VTG) is a protein produced by the liver of oviparous animals. It is being used as a... more Vitellogenin (VTG) is a protein produced by the liver of oviparous animals. It is being used as a biomarker for exposure to endocrine disruptors in many species. Rainbow trout Vtg has recently been sequenced by the conventional cDNA nucleotide approach. We focused on protein characterization of the intact protein and its derived tryptic and cyanogen bromide peptides by matrix-assisted laser desorption/ionisation and electrospray ionisation mass spectrometry. The molecular mass of the intact protein was found to be 183127 Da. A large number of unidentified peptide ions encourage further structural analysis to propose possible sequence variants and post-translational modifications.
Unusual ionization behavior was observed with novel antineoplastic curcumin analogues during the ... more Unusual ionization behavior was observed with novel antineoplastic curcumin analogues during the positive ion mode of matrix-assisted laser desorption ionization (MALDI) and dopant-free atmospheric pressure photoionization (APPI). The tested compounds produced an unusual significant peak designated as [M - H](+) ion along with the expected [M + H](+) species. In contrast, electrospray ionization, atmospheric pressure chemical ionization and the dopant-mediated APPI (dopant-APPI) showed only the expected [M + H](+) peak. The [M - H](+) ion was detected with all evaluated curcumin analogues including phosphoramidates, secondary amines, amides and mixed amines/amides. Our experiments revealed that photon energy triggers the ionization of the curcumin analogues even in the absence of any ionization enhancer such as matrix, solvent or dopant. The possible mechanisms for the formation of both [M - H](+) and [M + H](+) ions are discussed in this paper. In particular, three proposed mechanisms for the formation of [M - H](+) were evaluated. The first mechanism involves the loss of H2 from the protonated [M + H](+) species. The other two mechanisms include hydrogen transfer from the analyte radical cation or hydride abstraction from the neutral analyte molecule.
Vitellogenin is a protein produced by the liver of oviparous animals in response to circulating e... more Vitellogenin is a protein produced by the liver of oviparous animals in response to circulating estrogens. The amino acid sequence of vitellogenin from Atlantic cod (Gadus morhua) has not yet been determined. In this study Atlantic cod vitellogenin was characterized using a 'bottom-up' mass spectrometric approach. Vitellogenin synthesis was induced 'in vivo' with beta-estradiol, and subjected to trypsin digestion for characterization by matrix-assisted laser desorption/ionization quadrupole time-of-flight tandem mass spectrometry. A peptide mass fingerprint was obtained and 'de novo' sequencing of the most abundant tryptic peptides was performed by low-energy collision-induced dissociation tandem mass spectrometry. As a result of these experiments, the sequences of various tryptic peptides have been elucidated. The database search has shown that Atlantic cod vitellogenin shares a series of common peptides with the two different known vitellogenin sequences of haddock, a closely related species. These findings allow us to propose that Atlantic cod might also co-express at least two distinct forms of vitellogenin.
The molecular structure of the mutant form of the lipopolysaccharide of Aeromonas salmonicida was... more The molecular structure of the mutant form of the lipopolysaccharide of Aeromonas salmonicida was determined to contain an O-4 phosphorylated and O-5 substituted Kdo reducing group, and is proposed as the following: [molecular structure: see text] It was established that during the cleavage of this LPS with 1% acetic acid, to release the core oligosaccharide from the Lipid A portion, we obtained a degraded core oligosaccharide which eliminated its phosphate group with extreme facility. The precise molecular structure of this dephosphorylated core was deduced by electrospray mass spectrometry and is proposed as the following:[molecular structure: see text] Low energy collision ESI-QqTOF-MS/MS analysis of the dephosphorylated core oligosaccharide confirmed the presence of the O-5 glycosylated 4,8- and 4,7-anhydro derivatives of the enolizable alpha-keto-acids. The CID tandem mass spectrometric analysis of the heterogeneous mixture of the permethylated core oligosaccharide established the unreported methylation reaction on the diastereomeric 4,8- and 4,7-anhydro alpha-keto-acids and the complete permethylation and addition reaction of the O-5 glycosylated open chain reducing end terminal D-arabino-3-en-2-ulonic acid. The stereo-specific fragmentation routes obtained during the tandem mass spectrometric analysis permitted the precise sequencing of this dephosphorylated rough core oligosaccharide of the mutant LPS of A. salmonicida.
Electrospray ionization quadrupole time-of-flight (ESI-QqToF) mass spectra of the zwitteronic sal... more Electrospray ionization quadrupole time-of-flight (ESI-QqToF) mass spectra of the zwitteronic salts naloxonazine dihydrochloride 1 and naloxone hydrochloride 2, a common series of morphine opiate receptor antagonists, were recorded using different declustering potentials. The singly charged ion [M+H-2HCl](+) at m/z 651.3170 and the doubly charged ion [M+2H-2HCl](2+) at m/z 326.1700 were noted for naloxonazine dihydrochloride 1; and the singly charged ion [M+H-HCl](+) at m/z 328.1541 was observed for naloxone hydrochloride 2. Low-energy collision-induced dissociation tandem mass spectrometry (CID-MS/MS) experiments established the fragmentation routes of these compounds. In addition to the characteristic diagnostic product ions obtained, we noticed the formation of a series of radical product ions for the zwitteronic compounds 1 and 2, and also the formation of a distonic ion product formed from the singly charged ion [M+H-HCl](+) of naloxone hydrochloride 2. Confirmation of the various established fragmentation routes was effected by conducting a series of ESI-CID-QqTof-MS/MS product ion scans, which were initiated by CID in the atmospheric pressure/vacuum interface using a higher declustering potential. Deuterium labeling was also performed on the zwitteronic salts 1 and 2, in which the hydrogen atoms of the OH and NH groups were exchanged with deuterium atoms. Low-energy CID-QqTof-MS/MS product ion scans of the singly charged and doubly charged deuteriated molecules confirmed the initial fragmentation patterns proposed for the protonated molecules. Precursor ion scan analyses were also performed with a conventional quadrupole-hexapole-quadrupole tandem mass spectrometer and allowed the confirmation of the genesis of some diagnostic ions.
Recently, we identified a novel disulfide oxidoreductase, SdbA, in the oral bacterium Streptococc... more Recently, we identified a novel disulfide oxidoreductase, SdbA, in the oral bacterium Streptococcus gordonii. Disulfide oxidoreductases form disulfide bonds in nascent proteins using a CXXC catalytic motif. Typically, the N-terminal cysteine interacts with substrates, while the C-terminal cysteine is buried and only reacts with the first cysteine of the motif. In this study, we investigated the SdbA C(86) P(87) D(88) C(89) catalytic motif. In vitro, SdbA single cysteine variants at the N or C-terminal position (SdbAC86P and SdbAC89A ) were active, but displayed different susceptibility to oxidation, and N-terminal cysteine was prone to sulfenylation. In S. gordonii, mutants with a single N-terminal were inactive and formed unstable disulfide adducts with other proteins. Activity was partially restored by inactivation of pyruvate oxidase, a hydrogen peroxide generator. Presence of the C-terminal cysteine alone (in the SdbAC86P variant) could complement the ΔsdbA mutant and restore di...
NATO Science for Peace and Security Series A: Chemistry and Biology, 2010
... to 17b-estradiol in the granulose cells by the action of cytochrome P450 aromatase ... mRNA c... more ... to 17b-estradiol in the granulose cells by the action of cytochrome P450 aromatase ... mRNA can be qualitatively assayed by direct northern blotting while more sophis-ticated methods ... Vitellogenin, the egg yolk precursor protein, has become a popular biomarker for measuring ...
European journal of mass spectrometry (Chichester, England), 2003
Vitellogenin (VTG) is a protein produced by the liver of oviparous animals. It is being used as a... more Vitellogenin (VTG) is a protein produced by the liver of oviparous animals. It is being used as a biomarker for exposure to endocrine disruptors in many species. Rainbow trout Vtg has recently been sequenced by the conventional cDNA nucleotide approach. We focused on protein characterization of the intact protein and its derived tryptic and cyanogen bromide peptides by matrix-assisted laser desorption/ionisation and electrospray ionisation mass spectrometry. The molecular mass of the intact protein was found to be 183127 Da. A large number of unidentified peptide ions encourage further structural analysis to propose possible sequence variants and post-translational modifications.
Unusual ionization behavior was observed with novel antineoplastic curcumin analogues during the ... more Unusual ionization behavior was observed with novel antineoplastic curcumin analogues during the positive ion mode of matrix-assisted laser desorption ionization (MALDI) and dopant-free atmospheric pressure photoionization (APPI). The tested compounds produced an unusual significant peak designated as [M - H](+) ion along with the expected [M + H](+) species. In contrast, electrospray ionization, atmospheric pressure chemical ionization and the dopant-mediated APPI (dopant-APPI) showed only the expected [M + H](+) peak. The [M - H](+) ion was detected with all evaluated curcumin analogues including phosphoramidates, secondary amines, amides and mixed amines/amides. Our experiments revealed that photon energy triggers the ionization of the curcumin analogues even in the absence of any ionization enhancer such as matrix, solvent or dopant. The possible mechanisms for the formation of both [M - H](+) and [M + H](+) ions are discussed in this paper. In particular, three proposed mechanisms for the formation of [M - H](+) were evaluated. The first mechanism involves the loss of H2 from the protonated [M + H](+) species. The other two mechanisms include hydrogen transfer from the analyte radical cation or hydride abstraction from the neutral analyte molecule.
Vitellogenin is a protein produced by the liver of oviparous animals in response to circulating e... more Vitellogenin is a protein produced by the liver of oviparous animals in response to circulating estrogens. The amino acid sequence of vitellogenin from Atlantic cod (Gadus morhua) has not yet been determined. In this study Atlantic cod vitellogenin was characterized using a 'bottom-up' mass spectrometric approach. Vitellogenin synthesis was induced 'in vivo' with beta-estradiol, and subjected to trypsin digestion for characterization by matrix-assisted laser desorption/ionization quadrupole time-of-flight tandem mass spectrometry. A peptide mass fingerprint was obtained and 'de novo' sequencing of the most abundant tryptic peptides was performed by low-energy collision-induced dissociation tandem mass spectrometry. As a result of these experiments, the sequences of various tryptic peptides have been elucidated. The database search has shown that Atlantic cod vitellogenin shares a series of common peptides with the two different known vitellogenin sequences of haddock, a closely related species. These findings allow us to propose that Atlantic cod might also co-express at least two distinct forms of vitellogenin.
The molecular structure of the mutant form of the lipopolysaccharide of Aeromonas salmonicida was... more The molecular structure of the mutant form of the lipopolysaccharide of Aeromonas salmonicida was determined to contain an O-4 phosphorylated and O-5 substituted Kdo reducing group, and is proposed as the following: [molecular structure: see text] It was established that during the cleavage of this LPS with 1% acetic acid, to release the core oligosaccharide from the Lipid A portion, we obtained a degraded core oligosaccharide which eliminated its phosphate group with extreme facility. The precise molecular structure of this dephosphorylated core was deduced by electrospray mass spectrometry and is proposed as the following:[molecular structure: see text] Low energy collision ESI-QqTOF-MS/MS analysis of the dephosphorylated core oligosaccharide confirmed the presence of the O-5 glycosylated 4,8- and 4,7-anhydro derivatives of the enolizable alpha-keto-acids. The CID tandem mass spectrometric analysis of the heterogeneous mixture of the permethylated core oligosaccharide established the unreported methylation reaction on the diastereomeric 4,8- and 4,7-anhydro alpha-keto-acids and the complete permethylation and addition reaction of the O-5 glycosylated open chain reducing end terminal D-arabino-3-en-2-ulonic acid. The stereo-specific fragmentation routes obtained during the tandem mass spectrometric analysis permitted the precise sequencing of this dephosphorylated rough core oligosaccharide of the mutant LPS of A. salmonicida.
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