Kate L Henderson1, Mingjie Li1, Salette Martinez2, Edwin A Lewis1, Robert P Hausinger2, Joseph P Emerson3. 1. Department of Chemistry, Mississippi State University, Mississippi State, MS 39762, United States. 2. Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, MI 48824-4320, United States. 3. Department of Chemistry, Mississippi State University, Mississippi State, MS 39762, United States. Electronic address: jemerson@chemistry.msstate.edu.
Abstract
BACKGROUND: TauD is a nonheme iron(II) and α-ketoglutarate (αKG) dependent dioxygenase, and a member of a broader family of enzymes that oxidatively decarboxylate αKG to succinate and carbon dioxide thereby activating O2 to perform a range of oxidation reactions. However before O2 activation can occur, these enzymes bind both substrate and cofactor in an effective manner. Here the thermodynamics associated with substrate and cofactor binding to FeTauD are explored. METHODS: Thermal denaturation of TauD and its enzyme-taurine, enzyme-αKG, and enzyme-taurine-αKG complexes are explored using circular dichroism (CD) spectroscopy and differential scanning calorimetry (DSC). RESULTS: Taurine binding is endothermic (+26kcal/mol) and entropically driven that includes burial of hydrophobic surfaces to close the lid domain. Binding of αKG is enthalpically favorable and shows cooperativity with taurine binding, where the change in enthalpy associated with αKG binding (δΔHcal) increases from -30.1kcal/mol when binding to FeTauD to -65.2kcal/mol when binding to the FeTauD-taurine complex. CONCLUSIONS: The intermolecular interactions that govern taurine and αKG binding impact the global stability of TauD and its complexes, with clear and dramatic cooperativity between substrate and cofactor. GENERAL SIGNIFICANCE: Thermal denaturation of TauD and its enzyme-taurine, enzyme-αKG, and enzyme-taurine-αKG complexes each exhibited increased temperature stability over the free enzyme. Through deconvolution of the energetic profiles for all species studied, a thermodynamic cycle was generated that shows significant cooperativity between substrate and cofactor binding which continues to clarity the events leading up O2 activation.
BACKGROUND: TauD is a nonheme iron(II) and α-ketoglutarate (αKG) dependent dioxygenase, and a member of a broader family of enzymes that oxidatively decarboxylate αKG to succinate and carbon dioxide thereby activating O2 to perform a range of oxidation reactions. However before O2 activation can occur, these enzymes bind both substrate and cofactor in an effective manner. Here the thermodynamics associated with substrate and cofactor binding to FeTauD are explored. METHODS: Thermal denaturation of TauD and its enzyme-taurine, enzyme-αKG, and enzyme-taurine-αKG complexes are explored using circular dichroism (CD) spectroscopy and differential scanning calorimetry (DSC). RESULTS:Taurine binding is endothermic (+26kcal/mol) and entropically driven that includes burial of hydrophobic surfaces to close the lid domain. Binding of αKG is enthalpically favorable and shows cooperativity with taurine binding, where the change in enthalpy associated with αKG binding (δΔHcal) increases from -30.1kcal/mol when binding to FeTauD to -65.2kcal/mol when binding to the FeTauD-taurine complex. CONCLUSIONS: The intermolecular interactions that govern taurine and αKG binding impact the global stability of TauD and its complexes, with clear and dramatic cooperativity between substrate and cofactor. GENERAL SIGNIFICANCE: Thermal denaturation of TauD and its enzyme-taurine, enzyme-αKG, and enzyme-taurine-αKG complexes each exhibited increased temperature stability over the free enzyme. Through deconvolution of the energetic profiles for all species studied, a thermodynamic cycle was generated that shows significant cooperativity between substrate and cofactor binding which continues to clarity the events leading up O2 activation.
Authors: Jessica R O'Brien; David J Schuller; Victoria S Yang; Bret D Dillard; William N Lanzilotta Journal: Biochemistry Date: 2003-05-20 Impact factor: 3.162
Authors: Mingjie Li; Kate L Henderson; Salette Martinez; Robert P Hausinger; Joseph P Emerson Journal: J Biol Inorg Chem Date: 2018-06-19 Impact factor: 3.358