|
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, Newfoundland A1C 5S7, Canada. E-mail: banoubjo@dfo-
mpo.gc.ca or joebanoub@attglobal.net
Alejandro Cohen
Department of Biochemistry, Memorial University of Newfoundland, St. John’s, Newfoundland A1C 5S7,
Canada
Atef Mansour
Aquaculture Research Section, Science, Oceans and Environment Branch, Oceans Programs Division, Northwest Atlantic Fisheries Center, St
John’s, Newfoundland A1C 5X1, Canada
Pierre Thibault
Protein Analysis, Caprion Pharmaceuticals, 7150 Alexander Fleming, Montreal, Quebec H4S 2C8,
Canada |
|
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
effected using 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 used low-energy collisionally-induced dissociation (CID) in 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 14 tryptic peptides were sequenced and compared with the complete
rainbow trout VTG and the partial Atlantic salmon VTG sequences found in the Swiss-Prot database. De novo sequencing by CID MS/MS of 11 Atlantic salmon tryptic digest
peptides with 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 peptides, Atlantic salmon, mass spectrometry, de novo, sequencing |
