Top-down mass spectrometry reveals new sequence variants of the major bovine seminal plasma protein PDC-109.
The major protein of bovine seminal plasma, PDC-109, is a 109-residue polypeptide that exists as a polydisperse aggregate under native conditions. The oligomeric state of this aggregate varies with ionic strength and the presence of lipids. Binding of PDC-109 to choline phospholipids on the sperm plasma membrane results in an efflux of cholesterol and choline phospholipids, which is an important step in sperm capacitation. In this study, Fourier transform ion cyclotron resonance mass spectrometry was used to analyze PDC-109 purified from bovine seminal plasma. In addition to the previously known PDC-109 variants, four new sequence variants were identified by top-down mass spectrometry. For example, a protein variant containing point mutations P10L and G14R was identified along with another form having a 14-residue truncation in the N-terminal region. Two other minor variants could also be identified from the affinity-purified PDC-109. These results demonstrate that PDC-109 is naturally produced as a mixture of several protein forms, most of which have not been detected in previous studies. Native mass spectrometry revealed that PDC-109 is exclusively monomeric at low protein concentrations, suggesting that the protein oligomers are weakly bound and can easily be disrupted. Ligand binding to PDC-109 was also investigated, and it was observed that two molecules of O-phosphorylcholine bind to each PDC-109 monomer, consistent with previous reports.
Department of Chemistry, University of Eastern Finland, FI-80101, Joensuu, Finland.
Mikko
M
Laitaoja
Rajeshwer S
RS
Sankhala
Musti J
MJ
Swamy
Janne
J
Jänis
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D017354:Point Mutation; D012661:Semen / Q000737:chemistry*; D029607:Seminal Vesicle Secretory Proteins / Q000032:analysis / Q000737:chemistry* / Q000145:classification; D020539:Sequence Analysis, Protein; D017550:Spectroscopy, Fourier Transform Infrared
D029607:Seminal Vesicle Secretory Proteins; C424303:seminal vesicle secretory protein 109, Bos taurus