* To whom correspondence should be addressed.
Received June 1, 2017; Revision received August 9, 2017
Recombination of the isolated, fully reduced bd-type quinol oxidase from Escherichia coli with carbon monoxide was studied by pulsed absorption spectrophotometry with microsecond time resolution. Analysis of the kinetic phases of recombination was carried out using the global analysis of multiwavelength kinetic data (“Global fitting”). It was found that the unresolved photodissociation of CO is followed by a stepwise (with four phases) recombination with characteristic times (τ) of about 20 µs, 250 µs, 1.1 ms, and 24 ms. The 20-µs phase most likely reflects bimolecular recombination of CO with heme d. Two subsequent kinetic transitions, with τ ~ 250 µs and 1.1 ms, were resolved for the first time. It is assumed that the 250-µs phase is heterogeneous and includes two different processes: recombination of CO with ~7% of heme b595 and transition of heme d from a pentacoordinate to a transient hexacoordinate state in this enzyme population. The 24-ms transition probably reflects a return of heme d to the pentacoordinate state in the same protein fraction. The 1.1-ms phase can be explained by recombination of CO with ~15% of heme b558. Possible models of interaction of CO with different heme centers are discussed.
KEY WORDS: axial ligand, heme, cytochrome, respiratory chain, terminal oxidase, laser spectroscopy, flash photolysis, recombination, carbon monoxide