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Phosphorylated intermediates of two hepatic microsomal ATPases
The Biochemical Journal
|March 15, 1985
Summary
Researchers identified two distinct enzymes in liver microsomes responsible for Ca2+-ATPase and Mg2+-ATPase activities. Controlling free ion concentrations with CDTA is crucial for studying Ca2+-ATPase.
Area of Science:
- Biochemistry
- Cell Biology
Background:
- Hepatic microsomal fractions exhibit Ca2+- and Mg2+-dependent ATPase activities.
- Distinguishing between these activities is essential for understanding cellular energy metabolism and transport processes.
Purpose of the Study:
- To differentiate between the phosphoenzyme intermediates of hepatic microsomal Ca2+- and Mg2+-dependent ATPases.
- To elucidate the roles of specific chelators in studying these enzymatic activities.
Main Methods:
- Utilized chelators EGTA and CDTA (trans-cyclohexane-1,2-diamine-NNN'N'-tetra-acetic acid) to distinguish enzyme intermediates.
- Characterized phosphoenzyme intermediates based on their stability to hydroxylamine and base, and molecular weight (Mr).
Main Results:
- Identified a Ca2+-ATPase intermediate as a hydroxylamine-labile, base-labile 125,000-Mr phosphoprotein.
- Identified a Mg2+-ATPase intermediate as a hydroxylamine-stable, base-stable 30,000-Mr phosphoprotein.
- Demonstrated that CDTA abolishes Mg2+-ATPase phosphoenzyme formation and significantly reduces basal ATPase activity, indicating Mg2+ dependence.
Conclusions:
- The study demonstrates the existence of two separate and distinct enzymes responsible for Ca2+- and Mg2+-ATPase activities in hepatic microsomes.
- Controlling free ion concentrations with CDTA is recommended for obtaining more accurate data on microsomal Ca2+-ATPase activity.