title: Mass spectrometry-based sequencing and SRM-based quantitation of two novel vitellogenin isoforms in the leatherback sea turtle (Dermochelys coriacea).
abstract: No biomarker has yet been discovered to identify the reproductive status of the endangered leatherback sea turtle (Dermochelys coriacea). Although vitellogenin (VTG) could be used for this, its sequence is not known in D. coriacea and no quantitative assay has been carried out in this species to date. Using de novo sequencing-based proteomics, we unambiguously characterized sequences of two different VTG isoforms that we named Dc-VTG1 and Dc-VTG2. To our knowledge, this is the first clear evidence of different VTG isoforms and the structural characterization of derived yolk proteins in reptiles. This work illustrates how massive de novo sequencing can characterize novel sequences when working on "exotic" nonmodel species in which even nucleotide sequences are not available. We developed assays for absolute quantitation of these two isoforms using selected reaction monitoring (SRM) mass spectrometry, thus providing the first SRM assays developed specifically for a nonsequenced species. Plasma levels of Dc-VTG1 and Dc-VTG2 decreased as the nesting season proceeded, and were closely related to the increased levels of reproductive effort. The SRM assays developed here therefore provide an original and efficient approach for the reliable monitoring of reproduction cycles not only in D. coriacea, but potentially in other turtle species.
authors.affiliation: Département Sciences Analytiques, Université de Strasbourg, IPHC, 25 rue Becquerel, 67087 Strasbourg, France.
authors.forename: Marine I
authors.initials: MI
authors.lastname: Plumel
authors.forename: Thierry
authors.initials: T
authors.lastname: Wasselin
authors.forename: Virginie
authors.initials: V
authors.lastname: Plot
authors.forename: Jean-Marc
authors.initials: JM
authors.lastname: Strub
authors.forename: Alain
authors.initials: A
authors.lastname: Van Dorsselaer
authors.forename: Christine
authors.initials: C
authors.lastname: Carapito
authors.forename: Jean-Yves
authors.initials: JY
authors.lastname: Georges
authors.forename: Fabrice
authors.initials: F
authors.lastname: Bertile
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D005260:Female; D008969:Molecular Sequence Data; D009425:Nesting Behavior; D020033:Protein Isoforms / Q000737:chemistry; D040901:Proteomics; D030162:Reptilian Proteins / Q000097:blood / Q000737:chemistry*; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid; D053719:Tandem Mass Spectrometry; D014426:Turtles / Q000502:physiology*; D014819:Vitellogenins / Q000097:blood / Q000737:chemistry*
chemical_list: D020033:Protein Isoforms; D030162:Reptilian Proteins; D014819:Vitellogenins