Reversible steps in the reaction of aldehydes with bacterial luciferase intermediates

Abstract

Aliphatic aldehydes of different chain lengths were found to differ in their reaction at 22 °C with the B. harveyi luciferase peroxyflavin intermediate. Although similar quantum yields were obtained in the luciferase reaction with the different chain-length aldehydes, the catalytic turnover rates differed. The kinetics of a reaction utilizing two aldehydes of different chain lengths can thus indicate the degree to which the aldehyde reaction is reversible. By such criteria the reactions of octanal and decanal were found to be readily reversible, while that of dodecanal was not. This conclusion was supported both by the effects of long-chain alcohols, which are competitive inhibitors, and by the secondary addition of hydroxylamine, an aldehyde trapping agent. The results are consistent with a model in which there are many intermediates along the reaction path. Since the reactions are monitored by decay of luminescence intensity, it is difficult to determine the position of the rate-determining step. For octanal and decanal the rate-limiting step could be at an early reversible stage of the reaction, but later for dodecanal, subsequent to a less reversible step, but still prior to the final irreversible step which populates the excited state.