Presence of endogenous calcium ion and its functional and structural regulation in horseradish peroxidase.
The endogenous calcium ion (Ca2+) in horseradish peroxidase (HRP) was removed to cause substantial changes in the proton NMR spectra of the enzyme in various oxidation/spin states. The spectral changes were interpreted as arising from the substantial alterations in the heme environments, most likely the heme proximal and distal sides. The comparative kinetic and redox studies revealed that these conformational changes affect the reduction process of compound II, resulting in the decrease of the enzymatic activity of HRP. It is also revealed from the ESR spectrum and the temperature dependences of the NMR and optical absorption spectra of the Ca2+-free enzyme that the heme iron atom of the Ca2+-free enzyme is in a thermal spin mixing between ferric high and low spin states, in contrast to that of the native enzyme. These results show that Ca2+ functions in maintaining the protein structure in the heme environments as well as the spin state of the heme iron, in favor of the enzymatic activity of HRP.
D002118:Calcium / Q000502:physiology*; D004578:Electron Spin Resonance Spectroscopy; D006418:Heme / Q000378:metabolism; D006735:Horseradish Peroxidase / Q000378:metabolism*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D010084:Oxidation-Reduction; D010544:Peroxidases / Q000378:metabolism*; D011487:Protein Conformation; D013329:Structure-Activity Relationship
D006418:Heme; D006735:Horseradish Peroxidase; D010544:Peroxidases; D002118:Calcium