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Eur. J. Mass Spectrom. DOI: 10.1255/ejms.588

Characterization and de Novo Sequencing of Atlantic Salmon Vitellogenin Protein by Elctrospray Tandem and Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry

Joseph Banoub
Special Projects, Oceans Division, Northwest Atlantic Fisheries Center, St John's, Newfoundland, A1C 5X1, Canada and Department of Biochemistry, Memorial University of Newfoundland. St. John's, A1C 5S7. Canada
Alejandro Cohen
Department of Biochemistry, Memorial University of Newfoundland. St. John's, A1C 5S7. Canada
Pierre Thibault
Protein Analysis Caprion Pharmaceuticals 7150 Alexander Fleming Montreal, Quebec H4S 2C8
Atef Mansour
Special Projects, Oceans Division, Northwest Atlantic Fisheries Center, St John's, Newfoundland, A1C 5X1, Canada

ABSTRACT:
ABSTRACT: Vitellogenin (VTG) is a protein produced by the liver of oviparous animals in response to circulating estrogens. In the plasma of males and immature females, VTG is undetectable. VTG has been used as a biomarker for exposure to endocrine disruptors in many species. In the present study, characterization of intact Atlantic Salmon VTG was done by matrix-assisted laser desorption/ionization-Time-of-flight mass spectrometry (MALDI-TOF-MS). Tryptic digest peptides were analyzed by MALDI-TOF-MS to obtain a peptide mass fingerprint. De Novo sequencing of the tryptic peptides was performed by low energy collisionally induced dissociation (CID) using an electrospray ionization-quadrupole-TOF orthogonal hybrid mass spectrometer (ESI-Q-TOF-MS/MS). The interpretation of the product ion spectra obtained from the ESI-Q-TOF-MS/MS was done by Lutefisk, a computer based software algorithm. The molecular mass of the intact protein was found to be 187335 Da. A total of fourteen tryptic peptides were sequenced and compared to the complete Rainbow Trout VTG and to the partial Atlantic Salmon VTG sequences found in the Swiss-Prot database. De Novo sequencing by CID-MS/MS of eleven Atlantic Salmon tryptic digest peptides of selected precursor ions at m/z 788.24, 700.20, 794.75, 834.31, 889.28, 819.79, 865.27, 843.81, 572.20, 573.66 and 561.68 showed high homology with the known sequence of Rainbow Trout VTG. The last two precursor peptide ions, found at m/z 573.66 and m/z 561.68 also specifically matched the known portion of the Atlantic Salmon VTG sequence. Finally, three tryptic precursor peptide ions found at m/z 795.18, 893.28 and 791.05, provided product ion spectra, which were exclusive to the unsequenced portion of the Atlantic Salmon VTG

Keywords: Vitellogenin, Characterization, Ecotoxicology, Tryptic, Atlantic Salmon, Mass spectrometry, De Novo, Sequencing

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