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Determination of the complete covalent structure of the major glycoform of DQH sperm surface protein, a novel trypsin-resistant boar seminal plasma O-glycoprotein related to pB1 protein

Protein science. 1999 Jul ; 8 (7) : 1551-6.

Protein Sci
ISSN 0961-8368
Source

Language English Country United States Media print

Document type Journal Article, Research Support, Non-U.S. Gov't

Persistent link https://www.medvik.cz/link/pmid10422846

Online Full text

PubMed 10422846
PubMed Central PMC2144377
DOI 10.1110/ps.8.7.1551
Knihovny.cz E-resources

MeSH
Glycoproteins chemistry MeSH
Glycosylation MeSH
Protein Conformation MeSH
Membrane Glycoproteins chemistry MeSH
Molecular Sequence Data MeSH
Molecular Weight MeSH
Swine MeSH
Amino Acid Sequence MeSH
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization MeSH
Carrier Proteins chemistry MeSH
Animals MeSH
Check Tag
Male MeSH
Animals MeSH
Publication type
Journal Article MeSH
Research Support, Non-U.S. Gov't MeSH
Names of Substances
DQH sperm surface protein, Sus scrofa MeSH Browser
Glycoproteins MeSH
Membrane Glycoproteins MeSH
pAIF-1 protein, Sus scrofa MeSH Browser
Carrier Proteins MeSH

The complete covalent structure of a novel boar DQH sperm surface protein resistant to many classical procedures of enzymatic fragmentation was determined. The relative molecular mass of the major form of this protein determined by ESI-MS and MALDI-MS was 13,065.2+/-1.0 and 13,065.1, respectively. However, additional peaks differing by 162 Da (i.e., minus hexose), 365 Da (i.e., minus hexose and N-acetylhexosamine), 146 Da (i.e., plus deoxyhexose), and 291 Da (i.e., plus sialic acid) indicated the heterogeneity due to differences in glycosylation. The complete covalent structure of the protein was determined using automated Edman degradation, MALDI-MS, and post-source decay (PSD) MALDI-MS, and shown to consist of N-terminal O-glycosylated peptide followed by two fibronectin type II repeats. The carbohydrates are O-glycosidically linked to threonine 10, as confirmed by PSD MALDI-MS of the isolated N-terminal glycopeptide. Eight cysteine residues of the protein form four disulfide bridges, the positions of which were assigned from MALDI-MS and Edman degradation data. We conclude that mass spectral techniques provide an indispensable tool for the detailed analysis of the covalent structure of proteins, especially those that are refractory to standard approaches of protein chemistry.

Department of Biochemistry Faculty of Science Charles University Prague Praha Czech Republic

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