Protein binding in deactivation of ferrylmyoglobin by chlorogenate and ascorbate

Kinetics of reduction of iron(IV) in ferrylmyoglobin by chlorogenate in neu tral or moderately acidic aqueous solutions (0.16 M NaCl) to yield metmyogl obin was studied using stopped flow absorption spectroscopy. The reaction o ccurs by direct bimolecular electron transfer with (2.7 +/- 0.3) x 10(3) M- 1.s(-1) at 25.0 degrees C (Delta H-# = 59 +/- 6 kJ mol(-1), Delta S-# = 15 +/- 22 J.mol(-1) K-1) for protonated ferrylmyoglobin (pK(a) = 4.95) and wit h 216 +/- 50 M-1 s(-1) (Delta H-# = 73 +/- 8 kJ . mol(-1), Delta S-# = 41 /- 30 J.mol(-1).K-1) for nonprotonated ferrylmyoglobin in parallel with red uction of a chlorogenate/ferrylmyoglobin complex by a second chlorogenate m olecule with (8.6 +/- 1.1) x 10(2) M-1 s(-1) (Delta S-# = 74 +/- 8 kJ mol-l , Delta S-# = 59 +/- 28 J mol(-1) K-1) for protonated ferrylmyoglobin and w ith 61 +/- 9 M-1.s(-1) (Delta H-# = 82 +/- 12 kJ.mol(-1), Delta S-# = 63 +/ - 41 J mol(-1) K-1) for nonprotonated ferrylmyoglobin, Previously published data on ascorbate reduction of ferrylmyoglobin are reevaluated according t o a similar mechanism. For both protonated and nonprotonated ferrylmyoglobi n the binding constant of chlorogenate is similar to 300 M-1, and the modul ation of ferrylmyoglobin as an oxidant by chlorogenate (or ascorbate)leads to a novel antioxidant interaction for reduction of ferrylmyoglobin by asco rbate in mixtures with chlorogenate.