Human ovarian tumor-associated trypsin. Its purification and characterization from mucinous cyst fluid and identification as an activator of pro-urokinase.
In search of the target protease for the tumor-associated trypsin inhibitor TATI we recently identified a trypsin-like protease in cyst fluid of mucinous ovarian tumors (Stenman, U.-H., Koivunen, E., and Vuento, M. (1988) Biol. Chem. Hoppe-Seyler 369, 9-14). We have now purified this protease and demonstrate that it represents isoenzyme forms of trypsinogen, here called tumor-associated trypsin(ogen)s (TAT). The purification procedure comprised batchwise anion exchange chromatography, immunoaffinity chromatography with antibodies to trypsin, and separation of the two isoenzymes by reverse phase chromatography. In sodium dodecyl sulfate (SDS)-gel electrophoresis, the TAT-1 and TAT-2 isoenzymes have relative molecular weights (Mr) of 25,000 and 28,000, respectively, TAT-2 being the major component. The amino-terminal amino acid sequences correspond to those of pancreatic trypsinogen-1 and -2, respectively, and activation of the zymogens results in cleavage of a NH2-terminal activation peptide of 8 residues characteristic of trypsinogen. Isoelectric focusing in the presence of urea gives pI values of about 5 and 4 for TAT-1 and -2, respectively. The substrate specificities of the two TAT isoenzymes are very similar to, but not identical with, those of trypsin-1 and trypsin-2, respectively, suggesting slight differences in substrate binding site. TAT was found to be an efficient activator of pro-urokinase. Hence, TAT could take part in the protease cascade associated with tumor invasion.
Koivunen
E
E
Department I of Obstetrics and Gynecology, Helsinki University Central Hospital, Finland.
Huhtala
M L
ML
Stenman
U H
UH
D000595:Amino Acid Sequence; D003537:Cystadenoma / Q000201:enzymology*; D004792:Enzyme Precursors / Q000302:isolation & purification / Q000378:metabolism; D005260:Female; D006801:Humans; D007527:Isoenzymes / Q000302:isolation & purification / Q000502:physiology; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000302:isolation & purification*; D010048:Ovarian Cysts / Q000201:enzymology*; D010051:Ovarian Neoplasms / Q000201:enzymology*; D010179:Pancreas / Q000201:enzymology; D010960:Plasminogen Activators / Q000302:isolation & purification* / Q000502:physiology; D014357:Trypsin / Q000302:isolation & purification*; D014568:Urokinase-Type Plasminogen Activator / Q000302:isolation & purification* / Q000502:physiology
D004792:Enzyme Precursors; D007527:Isoenzymes; D009363:Neoplasm Proteins; D010960:Plasminogen Activators; D014357:Trypsin; D014568:Urokinase-Type Plasminogen Activator