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Related Experiment Videos

Energy changes during the formation and interconversion of enzyme-substrate complexes.

H Gutfreund, D R Trentham

    Ciba Foundation Symposium
    |January 1, 1975
    PubMed
    Summary

    Rapid reaction and isotope techniques reveal enzyme reaction mechanisms. Chemical catalysis steps are often fast with small free-energy changes, unlike substrate binding.

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    Rapid-flow techniques and their contributions to enzymology.

    Trends in biochemical sciences·1999

    Area of Science:

    • Biochemistry
    • Enzyme kinetics
    • Chemical kinetics

    Background:

    • Enzyme reactions involve multiple steps, including substrate binding, chemical catalysis, and product release.
    • Understanding the kinetics of individual steps is crucial for elucidating enzyme mechanisms.
    • The Haldane relation provides a framework but lacks detailed mechanistic insights.

    Purpose of the Study:

    • To determine rate and equilibrium constants for individual steps in enzyme reactions.
    • To complement the Haldane relation with detailed reaction mechanism information.
    • To investigate three specific enzyme reactions using advanced kinetic techniques.

    Main Methods:

    • Rapid reaction methods
    • Isotope techniques
    • Absorption spectroscopy
    • Fluorescence spectroscopy
    • Chemical quenching

    Main Results:

    • Cardiac lactate dehydrogenase (EC 1.1.1.27): Chemical catalysis is fast, equilibrium constant near unity; free energy changes reflect hydrogen transfer.
    • Triosephosphate isomerase (EC 5.3.1.1): Enzyme-bound substrate-to-product interconversion rate is comparable to product dissociation.
    • Myosin subfragment 1 with ATP: Chemical step equilibrium constant is 9; ATP binding involves a large free-energy change.

    Conclusions:

    • Enzyme catalysis steps are often rapid with small free-energy changes compared to binding/dissociation.
    • Kinetic parameters of elementary steps provide mechanistic details beyond the Haldane relation.
    • Findings have implications for energy transduction in muscle contraction and ATP biosynthesis.

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