Photothermal studies of CO photodissociation from mixed valence Escherichia coli cytochrome bo₃

Here, we report the volume and enthalpy changes accompanying CO photodissociation from the mixed valence form of cytochrome bo₃ oxidase from Escherichia coli. The results of photoacoustic calorimetry indicate two kinetic phases with distinct volume and enthalpy changes accompanying CO photodissociation from heme o₃ and its transfer to Cu_B. The first phase occurring on a timescale of <50 ns is characterized by a volume decrease of -1.3 ± 0.3 mL mol^-1 and enthalpy change of 32 ± 1.6 kcal mol^-1. Subsequently, a volume increase of 2.9 mL mol^-1 with an enthalpy change of -5.3 ± 2.5 kcal mol ^-1 is observed with the lifetime of ∼250 ns (this phase has not been detected in previous optical studies). These volume and enthalpy changes differ from the volume and enthalpy changes observed for CO dissociation from fully reduced cytochrome bo₃ oxidase indicating that the heme o ₃/Cu_B active site dynamics are affected by the redox state of heme b.