Localization of the disulfide bonds in the NH2-terminal domain of the cellular receptor for human urokinase-type plasminogen activator. A domain structure belonging to a novel superfamily of glycolipid-anchored membrane proteins.
The receptor for human urokinase-type plasminogen activator (uPAR) is synthesized as a 313-residue-long polypeptide containing 28 cysteine residues, the pattern of which defines three homologous repeats within the protein. These entities are believed to represent a novel protein domain structure, of which the NH2-terminal domain of uPAR can be covalently cross-linked to the epidermal growth factor-like module of urokinase after receptor-ligand interaction. The NH2-terminal domain of a recombinant, soluble uPAR derivative, labeled with [35S]cysteine, was isolated after limited proteolysis with chymotrypsin. The four disulfide bonds present within this domain were assigned by a combination of plasma desorption mass spectrometry, amino acid composition, and sequence analyses of peptides generated by trypsin, endoproteinase Asp-N, and thermolysin. The following disulfide bond structure was determined: Cys3-Cys24, Cys6-Cys12, Cys17-Cys45, and Cys71-Cys76. Similar cysteine pairing is likely to be found within other members of this protein superfamily, i.e. the membrane inhibitor of reactive lysis, Ly-6, and the remaining two domains of uPAR. However, an additional pair of cysteines present within these domains probably forms a fifth disulfide bond.
Finsen Laboratory, Rigshospitalet Opg 86.21, Copenhagen, Denmark.
M
M
Ploug
M
M
Kjalke
E
E
Rønne
U
U
Weidle
G
G
Høyer-Hansen
K
K
Danø
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D002918:Chymotrypsin; D006224:Cricetinae; D004220:Disulfides / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel; D017261:Glycosylphosphatidylinositols / Q000737:chemistry*; D006801:Humans; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010960:Plasminogen Activators; D011956:Receptors, Cell Surface / Q000737:chemistry*; D055293:Receptors, Urokinase Plasminogen Activator; D013820:Thermolysin; D014357:Trypsin; D014568:Urokinase-Type Plasminogen Activator
D004220:Disulfides; D017261:Glycosylphosphatidylinositols; C524898:PLAUR protein, human; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D010960:Plasminogen Activators; D002918:Chymotrypsin; D014357:Trypsin; D014568:Urokinase-Type Plasminogen Activator; D013820:Thermolysin