Characterization and De Novo Sequencing of Atlantic Salmon Vitellogenin Protein by Electrospray Tandem and Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry
peer-reviewed · European Journal of Mass Spectrometry · 2004
| Date | 2004-02-01 |
| Type | peer-reviewed |
| Venue | European Journal of Mass Spectrometry |
| Publisher | SAGE Publications |
| Contribution | downstream-application |
| DOI | 10.1255/ejms.588 |
| Citations (OpenAlex) | 21 |
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 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.
Methods and tools
- Atlantic salmon vitellogenin sequencing: Characterised salmon vitellogenin by MALDI and Q-ToF MS/MS, de novo sequencing 14 tryptic peptides with Lutefisk, three of them from a portion of the protein not yet in databases.
Methods it uses
- Lutefisk: First widely-used heuristic de novo tool