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Characterization of a novel calcium-binding 90-kDa glycoprotein (BM-90) shared by basement membranes and serum
1Max-Planck-Institut für Biochemie, Martinsried, Federal Republic of Germany.
Abstract:
The protein BM-90 was solubilized from the mouse Engelbreth-Holm-Swarm tumor with neutral buffers in molar yields lower (15-30%) than found for other basement membrane proteins (e.g. laminin, BM-40). The purified protein was shown to be rich in cysteine (5 mol%) and to change in SDS electrophoresis from an 84-kDa position to a 95-kDa one upon reduction. BM-90 was also shown to be a calcium-binding protein. The N-terminal sequence of BM-90, as well as those of several internal peptides, showed no identity with any known protein sequences, indicating that it is a new protein. Specific radioimmunoassays showed no or only minor cross-reactions with other known basement membrane proteins. Immunological assays demonstrated BM-90 to be present in neutral salt extracts from mouse heart and kidney, in serum (20-40 micrograms/ml) and in the medium of various cultured cells (0.1-1 microgram/ml). The protein in these samples was identical in size to BM-90 purified from the tumor, indicating that negligible degradation occurs during purification. An extracellular matrix localization of BM-90 was shown by immunofluorescence for Reichert's membrane, lens capsules and other basement membranes. Thus, BM-90 appears to be a novel basement membrane protein whose functions remain to be studied.
Insights
Researchers identified a novel protein, BM-90, in mouse tumors. This cysteine-rich, calcium-binding protein is present in various tissues and extracellular matrices, suggesting a new role in basement membranes.
Area of Science:
- Biochemistry
- Cell Biology
- Proteomics
Background:
- Basement membranes are crucial extracellular matrices involved in tissue structure and cell signaling.
- Several basement membrane proteins like laminin and BM-40 have been characterized.
- The Engelbreth-Holm-Swarm (EHS) mouse tumor is a rich source for basement membrane protein isolation.
Purpose of the Study:
- To isolate and characterize a novel protein from the mouse EHS tumor.
- To determine the biochemical properties and potential functions of this new protein.
- To investigate the presence and localization of the protein in various biological samples.
Main Methods:
- Protein solubilization from EHS tumor using neutral buffers.
- SDS-PAGE electrophoresis to determine molecular weight and reduction sensitivity.
- N-terminal sequencing and internal peptide analysis for protein identification.
- Calcium-binding assays.
- Specific radioimmunoassays and immunological assays.
- Immunofluorescence microscopy for extracellular matrix localization.
Main Results:
- A novel protein, BM-90, was purified from mouse EHS tumor with low molar yields (15-30%).
- BM-90 is rich in cysteine (5 mol%) and exhibits a molecular weight shift from 84 kDa to 95 kDa upon reduction.
- It is a calcium-binding protein with no sequence homology to known proteins.
- BM-90 was detected in mouse heart, kidney, serum, and cultured cell media, localizing to extracellular matrices.
Conclusions:
- BM-90 is a newly discovered basement membrane protein with unique biochemical characteristics.
- Its presence in various tissues and extracellular matrices suggests a significant biological role.
- Further research is needed to elucidate the specific functions of BM-90 in basement membranes.