A novel 500,000 Da, linear, single chain extracellular protein synthesized by several childhood tumors.
A high molecular weight extracellular protein has been purified from cell culture medium of Ewing's sarcoma cell lines, by high performance liquid chromatography and electroelution from SDS-PAGE electrophoresis. This protein has an apparent molecular mass of about 500,000 Da on SDS-PAGE. Immunoprecipitation studies with several extracellular matrix glycoproteins (laminin, fibronectin) specific antisera indicate it is a separate protein. Reduction of disulphide bonds with 2-ME or DTT fails to significantly alter its migration on SDS-PAGE gels, other than a slight apparent increase in molecular mass, indicating an apparent single polypeptide chain structure. The slightly greater mobility observed in unreduced gels suggests one or more regions of intrachain disulfide bonding. It is sensitive to pepsin and trypsin, but resistant to bacterial collagenase indicating that it does not contain collagenous domains. Metabolic labelling with 3H-proline, 3H-leucine, and 35S-methionine indicate that this protein is proline-poor, but leucine, and especially methionine, rich. Sodium 35S-sulfate incorporation is totally negative and treatment with glycosaminoglycan degrading enzymes has no effect on the mobility of the protein on gels, unlike typical proteoglycans. This protein appears by rotary shadowing electron microscopy as a long, thin, filamentous molecule at least 500 nm (0.5 um) in length. The tissue localization and function are unknown at this time, but are under active investigation.
Scarpa
S
S
Dipartimento di Medicina Sperimentale, Università La Sapienza, Roma.
Reddy
M P
MP
Bressani
N
N
Mims
S
S
Massari
F
F
Triche
T J
TJ
D002648:Child; D003470:Culture Media; D005109:Extracellular Matrix / Q000378:metabolism*; D006801:Humans; D008854:Microscopy, Electron; D008970:Molecular Weight; D009363:Neoplasm Proteins / Q000096:biosynthesis* / Q000648:ultrastructure; D014407:Tumor Cells, Cultured / Q000378:metabolism*
D003470:Culture Media; D009363:Neoplasm Proteins