Related Experiment Videos
DNA methyltransferase polypeptides in mouse and human cells
Abstract:
DNA methyltransferase was isolated as a single polypeptide of 190 kDa from mouse P815 mastocytoma cells by immunoaffinity chromatography. This polypeptide seems to be highly susceptible to proteolytic degradation resulting in additional polypeptides in the size range of 150 to 190 kDa. A polypeptide of 190 kDa was immunoprecipitated by monoclonal anti-DNA methyltransferase antibodies from extracts of two different human cell lines, Raji and K562. The 190 kDa polypeptide was synthesized in rapidly proliferating cells and, albeit at a much lower rate, also in cells grown to saturating density. DNA methyltransferase polypeptides smaller than 190 kDa were synthesized neither in log phase nor in stationary phase cells.
Insights
Researchers isolated a 190 kDa DNA methyltransferase polypeptide from mouse cells using immunoaffinity chromatography. This key enzyme was also found in human cell lines and synthesized in both proliferating and non-proliferating cells.
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- DNA methyltransferase (DNMT) is crucial for epigenetic regulation.
- Understanding DNMT's structure and synthesis is vital for comprehending gene expression control.
Purpose of the Study:
- To isolate and characterize the DNA methyltransferase polypeptide.
- To investigate the synthesis and presence of DNA methyltransferase in different cellular conditions.
Main Methods:
- Immunoaffinity chromatography using monoclonal anti-DNA methyltransferase antibodies.
- Immunoprecipitation from mouse P815 mastocytoma cells and human cell lines (Raji, K562).
- Analysis of polypeptide sizes and synthesis rates in proliferating and stationary phase cells.
Main Results:
- A 190 kDa DNA methyltransferase polypeptide was successfully isolated from mouse cells.
- The 190 kDa polypeptide was also identified in human Raji and K562 cell lines.
- This 190 kDa polypeptide is synthesized in both rapidly proliferating and, at lower rates, in stationary phase cells, while smaller DNMT polypeptides are not synthesized under either condition.
Conclusions:
- The primary form of DNA methyltransferase is a 190 kDa polypeptide.
- This polypeptide is conserved across mouse and human cell lines.
- DNMT synthesis is regulated by cell proliferation status, with the 190 kDa form being the predominant species synthesized.