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Mouse DNA methylase. Intracellular location and degradation
R L Adams1, J Hill, J M McGarvey
1Department of Biochemistry, University of Glasgow, UK.
Cell Biophysics
|August 1, 1989
Summary
Stepwise trypsin treatment of mouse DNA methylase reveals its domain structure through nicking and fragment release. This process enhances enzyme activity and prevents aggregation, particularly in liver nuclear extracts.
Area of Science:
- Biochemistry
- Molecular Biology
- Enzymology
Background:
- DNA methylase plays a crucial role in epigenetic regulation.
- Understanding the structural and functional properties of DNA methylase is essential for comprehending gene expression control.
Purpose of the Study:
- To investigate the structural organization and domain architecture of mouse DNA methylase.
- To analyze the effects of enzymatic degradation on DNA methylase activity and stability.
Main Methods:
- Stepwise degradation of purified DNA methylase using trypsin.
- Analysis of enzyme fragments by denaturation and size determination (kD).
- Comparison of enzyme preparations from ascites cell nuclei and liver nuclei.
Main Results:
- Trypsin treatment introduced nicks into DNA methylase, releasing specific fragments (14, 18, 24, 28 kD), indicating a domain structure.
- Liver nuclear extracts contained pre-nicked enzyme with a major 100 kD fragment, unlike ascites extracts.
- Newborn mouse liver had more undegraded, nuclear-bound enzyme.
- Trypsin treatment boosted de novo activity and prevented aggregation.
Conclusions:
- Mouse DNA methylase exhibits a distinct domain structure revealed by limited proteolysis.
- Nuclear DNA methylase exists in different forms, with liver extracts showing more degradation.
- Enzymatic nicking can enhance DNA methylase activity and improve its handling properties.