Time-resolved infrared studies of the dynamics of ligand binding to cytochrome c oxidase

Time-resolved infrared spectroscopy (TRIRS) has been employed to study the reactions of small molecules with the cytochrome a3-CuB site of cytochrome c oxidase (CcO). All phases of these reactions have been investigated, from ultrafast phenomena (hundreds of femtoseconds) to relatively slow processes (milliseconds). The ligation dynamics immediately following photodissociation have been studied using a TRIRS technique with time resolution of less than 1 ps. The rate of photoinitiated transfer of CO from Fea32+ to CuB+ was measured directly by monitoring the development of the transient CuB+-CO absorption. The development of a stationary CuB+ spectrum which is constant until the CO dissociates from CuB+ occurs in less than 1 ps, indicating that the photoinitiated transfer of CO is remarkably fast. This unprecedented ligand transfer rate has profound implications with regard to the structure and dynamics of the cytochrome a3-CuB site, the functional architecture of the protein and coordination dynamics in general. The photodissociation and recombination of CN- has also been studied using a real-time TRIR technique. The CN- recombination rate of 430 s-1 is consistent with a recombination pathway similar to the one we have previously proposed for CO, in which a long-lived barrier to recombination is formed by the binding of an endogenous ligand L to Fea32+. The authors suggest the rate determining step for CN- recombination is the thermal dissociation of the Fea32+-L bond.