Posttranslational modifications of microfibril associated glycoprotein-1 (MAGP-1).
Microfibril-associated glycoprotein-1 (MAGP-1) is a small molecular weight protein associated with extracellular matrix microfibrils. Biochemical studies have shown that MAGP-1 undergoes several posttranslational modifications that may influence its associations with other microfibrillar components. To identify the sites in the molecule where posttranslational modifications occur, we expressed MAGP-1 constructs containing various point mutations as well as front and back half truncations in CHO cells. Characterization of transiently expressed protein showed that MAGP-1 undergoes O-linked glycosylation and tyrosine sulfation at sites in its amino-terminal half. This region of the protein also served as a major amine acceptor site for transglutaminase and mediated self-assembly into high molecular weight multimers through a glutamine-rich sequence. Fine mapping of the modification sites through mutational analysis demonstrated that Gln20 is a major amine acceptor site for the transglutaminase reaction and confirmed that a canonical tyrosine sulfation consensus sequence is the site of MAGP-1 sulfation. Our results also show that O-glycosylation occurs at more than one site in the molecule.
Trask
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BC
Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Broekelmann
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T
Ritty
T M
TM
Trask
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TM
Tisdale
C
C
Mecham
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RP
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D016466:CHO Cells; D002236:Carbohydrate Conformation; D002417:Cattle; D003285:Contractile Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D006224:Cricetinae; D005109:Extracellular Matrix / Q000235:genetics / Q000378:metabolism*; D016326:Extracellular Matrix Proteins; D005822:Genetic Vectors / Q000378:metabolism; D005973:Glutamine / Q000235:genetics; D006031:Glycosylation; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D011499:Protein Processing, Post-Translational / Q000235:genetics; D000072260:RNA Splicing Factors; D011994:Recombinant Proteins / Q000378:metabolism; D013431:Sulfates / Q000378:metabolism; D014162:Transfection; D011503:Transglutaminases / Q000235:genetics / Q000378:metabolism; D014333:Tropoelastin / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism
D003285:Contractile Proteins; D016326:Extracellular Matrix Proteins; D010446:Peptide Fragments; D000072260:RNA Splicing Factors; D011994:Recombinant Proteins; D013431:Sulfates; D014333:Tropoelastin; C016598:microfibrillar protein; D005973:Glutamine; D014443:Tyrosine; D011503:Transglutaminases