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Simultaneous Measurement of Superoxide/Hydrogen Peroxide and NADH Production by Flavin-containing Mitochondrial Dehydrogenases
Published on: February 24, 2018
Metal-Induced reversible structural interconversion of human mitochondrial NAD(P)+-dependent malic enzyme
Chu-Wei Kuo1, Hui-Chih Hung, Liang Tong
1Institute of Biochemistry, National Defense Medical Center, Taipei, Taiwan.
Proteins
|January 30, 2004
Summary
Lutetium (Lu3+) binding to human mitochondrial malic enzyme causes a slow, reversible structural change, leading to enzyme inhibition. Manganese (Mn2+) can reverse this inhibition by restoring the enzyme's active conformation.
Area of Science:
- Biochemistry
- Enzyme kinetics
- Structural biology
Background:
- Human mitochondrial NAD(P)+-dependent malic enzyme plays a crucial role in cellular metabolism.
- Previous studies indicated structural changes in the enzyme upon Lu3+ binding.
Purpose of the Study:
- To characterize the reversible slow-binding inhibition mechanism of Lu3+ on human mitochondrial malic enzyme.
- To investigate the structural interconversion between Mn2+- and Lu3+-bound forms of the enzyme.
Main Methods:
- Enzyme activity assays to determine kinetic parameters and inhibition constants.
- Fluorescence quenching experiments to probe conformational changes.
- Analysis of hyperbolic dependence of rate constants on inhibitor concentration.
Main Results:
- Lu3+ acts as a slow-binding inhibitor, with higher affinity for the isomerized enzyme form.
- The Lu3+-induced inhibition is reversible, with Mn2+ capable of reactivating the enzyme.
- Fluorescence quenching data confirm the reversibility of Lu3+-induced structural changes, while Mn2+ binding does not induce significant conformational changes.
Conclusions:
- Lu3+ binding induces a reversible isomerization of human mitochondrial malic enzyme to a catalytically inactive state.
- The slow kinetics of inhibition and reactivation are attributed to a structural rearrangement.
- Mn2+ can displace Lu3+ and restore the active conformation, highlighting the dynamic nature of the enzyme's metal-binding site.
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