Characterization of site-specific O-glycan structures within the mucin-like domain of alpha-dystroglycan from human skeletal muscle.
The glycosylation of the extracellular protein alpha-dystroglycan is important for its ligand-binding activity, and altered or blocked glycosylation is associated with several forms of congenital muscular dystrophies. By immunoprecipitation and sialic acid capture-and-release enrichment strategies, we isolated tryptic glycopeptides of alpha-dystroglycan from human skeletal muscle. Nano-liquid chromatography tandem mass spectrometry was used to identify both glycopeptides and peptides corresponding to the mucin-like and C-terminal domain of alpha-dystroglycan. The O-glycans found had either Hex-O-Thr or HexNAc-O-Ser/Thr anchored structures, which were often elongated and frequently, but not always, terminated with sialic acid. The HexNAc-O-Ser/Thr, but not Hex-O-Thr glycopeptides, displayed heterogeneity regarding glycan core structures and level of glycosylation site occupancy. We demonstrate for the first time glycan attachment sites of the NeuAcHexHexNAcHex-O structure corresponding to the anticipated Neu5Acalpha3Galbeta4GlcNAcbeta2Man-O-glycan (sLacNAc-Man), within the mucin-like domain of human alpha-dystroglycan from human skeletal muscle. Twenty-five glycopeptides were characterized from human alpha-dystroglycan, which provide insight to the complex in vivo O-glycosylation of alpha-dystroglycan.
Institute of Biomedicine, Department of Clinical Chemistry and Transfusion Medicine, Sahlgrenska Academy at the University of Gothenburg, Gothenburg, Sweden.
Johanna
J
Nilsson
Jonas
J
Nilsson
Göran
G
Larson
Ammi
A
Grahn
D000595:Amino Acid Sequence; D002240:Carbohydrate Sequence; D020134:Catalytic Domain; D049030:Dystroglycans / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D009077:Mucins / Q000737:chemistry*; D018482:Muscle, Skeletal / Q000737:chemistry / Q000378:metabolism*; D019158:N-Acetylneuraminic Acid / Q000737:chemistry / Q000378:metabolism; D010449:Peptide Mapping; D011134:Polysaccharides / Q000737:chemistry / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary / Q000502:physiology; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
C483081:DAG1 protein, human; D009077:Mucins; D011134:Polysaccharides; D049030:Dystroglycans; D019158:N-Acetylneuraminic Acid