Mass spectrometric characterization of full-length rat selenoprotein P and three isoforms shortened at the C terminus. Evidence that three UGA codons in the mRNA open reading frame have alternative functions of specifying selenocysteine insertion or translation termination.
Selenoprotein P is an abundant extracellular glycoprotein. Its mRNA contains 10 UGAs in an open reading frame terminated by a UAA. This predicts that full-length selenoprotein P will contain 10 selenocysteine residues. Full-length selenoprotein P and three smaller isoforms that have identical N termini have been demonstrated. Selenoprotein P was purified from rat plasma, and the four isoforms were separated by heparin chromatography and SDS-PAGE. Mass spectrometric peptide analysis of the full-length isoform verified 357 of its 366 predicted amino acid residues, including its C terminus and all 10 selenocysteines. The C termini of the smaller isoforms were characterized by mass spectrometry. The shortened isoforms terminated where the second, third, and seventh selenocysteine residues were predicted to be. This suggests that all isoforms arise from the same mRNA and that the UGAs that specify the second, third, and seventh selenocysteines in full-length selenoprotein P can alternatively serve to terminate translation, producing the shorter isoforms.
Ma
Shuguang
S
Mass Spectrometry Research Center, Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN 37232-2279, USA.
Hill
Kristina E
KE
Caprioli
Richard M
RM
Burk
Raymond F
RF
D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D018388:Codon, Terminator; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010449:Peptide Mapping; D014176:Protein Biosynthesis; D020033:Protein Isoforms / Q000737:chemistry* / Q000235:genetics; D011506:Proteins / Q000737:chemistry* / Q000235:genetics; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics; D051381:Rats; D017279:Selenocysteine / Q000737:chemistry*; D051149:Selenoprotein P; D051140:Selenoproteins; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D018388:Codon, Terminator; D020033:Protein Isoforms; D011506:Proteins; D012333:RNA, Messenger; D051149:Selenoprotein P; D051140:Selenoproteins; D017279:Selenocysteine