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A basement membrane-associated glycoprotein from skeletal muscle
Abstract:
We have isolated a major glycoprotein that appears to be associated with rat skeletal muscle basement membrane. We determined that the glycoprotein was part of the muscle cell surface complex when we found it to be enriched in preparations of muscle ghosts. We isolate the glycoprotein from homogenized muscle preextracted with 4 M and 8 M urea. It elutes as a major component in the presence of 8 M urea/50 mM 2-mercaptoethanol. Its apparent molecular weight on sodium dodecyl sulfate gels is 130,000. Amino acid analysis indicates that it is not a collagen but that it does contain small amounts of hydroxyproline and hydroxylysine. There may be collagenous domains in the glycoprotein molecule, for it is cleaved into three fragments by purified bacterial collagenase. Immunoperoxidase staining confirms that the 130,000-dalton protein is localized at the surface of adult skeletal muscle cells. It is probably a general basement membrane-associated glycoprotein because we found material immunologically cross-reactive with the muscle glycoprotein in basement membrane regions of kidney, liver, brain, and small intestine. We have shown the glycoprotein to be distinct from fibronectin, laminin, and types I, III, IV, and V collagens in enzyme-linked immunosorbent assays.
Insights
Researchers isolated a novel glycoprotein from rat skeletal muscle basement membrane. This 130,000-dalton protein is associated with muscle cell surfaces and found in other basement membranes.
Area of Science:
- Biochemistry
- Cell Biology
- Extracellular Matrix Research
Background:
- The skeletal muscle basement membrane is a critical component of muscle tissue.
- Understanding its associated proteins is essential for comprehending muscle structure and function.
Purpose of the Study:
- To isolate and characterize a major glycoprotein associated with rat skeletal muscle basement membrane.
- To determine its cellular localization and potential role within the basement membrane complex.
Main Methods:
- Isolation of glycoprotein from homogenized muscle using urea extraction.
- Electrophoretic analysis (SDS-PAGE) to determine molecular weight.
- Amino acid analysis and enzymatic digestion (collagenase) to assess protein composition.
- Immunoperoxidase staining for cellular localization.
- Enzyme-linked immunosorbent assays (ELISAs) to compare with known matrix proteins.
Main Results:
- A major glycoprotein (130,000 daltons) was isolated from rat skeletal muscle basement membrane.
- The glycoprotein is enriched in muscle cell surface preparations.
- Amino acid analysis revealed hydroxyproline and hydroxylysine, suggesting potential collagenous domains.
- Immunoperoxidase staining confirmed localization on adult skeletal muscle cell surfaces.
- Immunological cross-reactivity was observed in basement membranes of kidney, liver, brain, and small intestine.
- The glycoprotein was distinguished from fibronectin, laminin, and collagens types I, III, IV, and V.
Conclusions:
- A novel 130,000-dalton glycoprotein is a significant component of rat skeletal muscle basement membrane.
- This glycoprotein is localized to the muscle cell surface and is likely a general basement membrane-associated protein.
- It represents a distinct molecular entity, different from previously characterized matrix proteins like fibronectin and laminin.