Isolation and characterization of type IV procollagen, laminin, and heparan sulfate proteoglycan from the EHS sarcoma.
We have studied the extractability of type IV collagen, laminin, and heparan sulfate proteoglycan from EHS tumor tissue growth in normal and lathyritic animals. Laminin and heparan sulfate proteoglycan were readily extracted with chaotropic solvents from both normal and lathyritic tissue. The collagenous component was only solubilized from lathyritic tissue in the presence of a reducing agent. These results indicate that lysine-derived cross-links and disulfide bonds stabilize the collagenous component in the matrix but not the laminin or the heparan sulfate proteoglycan. The majority of the collagen present in the extracts had a native triple helix based upon the pattern of peptides resistant to pepsin digestion and visualization in the electron microscope by the rotary shadow technique. This protein was composed of chains (Mr 185000 and 170000) identical in migration to the chains of newly synthesized type IV procollagen. This finding confirms earlier work that indicates that the biosynthetic form, type IV procollagen, is incorporated as such in the basement membrane matrix. Material with smaller chains (Mr 160000 and 140000) appeared on storage in acetic acid solutions. These results indicate that the lower molecular weight collagen in acid extracts of basement membrane arises artifactually due to an endogenous acid-active protease.
D000818:Animals; D011508:Chondroitin Sulfate Proteoglycans / Q000302:isolation & purification*; D004229:Dithiothreitol; D006023:Glycoproteins / Q000302:isolation & purification*; D006025:Glycosaminoglycans / Q000302:isolation & purification*; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate / Q000302:isolation & purification*; D007797:Laminin; D007842:Lathyrism / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008854:Microscopy, Electron; D008970:Molecular Weight; D009363:Neoplasm Proteins / Q000302:isolation & purification*; D011347:Procollagen / Q000302:isolation & purification*; D011509:Proteoglycans / Q000302:isolation & purification*; D012513:Sarcoma, Experimental / Q000032:analysis*
D011508:Chondroitin Sulfate Proteoglycans; D004229:Dithiothreitol; D006023:Glycoproteins; D006025:Glycosaminoglycans; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate; D007797:Laminin; D009363:Neoplasm Proteins; D011347:Procollagen; D011509:Proteoglycans