Halide dependence of the halorhodopsin photocycle as measured by time-resolved infrared spectra

Time-resolved Fourier transform infrared (FTIR) difference spectra of the halorhodopsin (hR) photocycle have been collected from 3μs to 100 ms in saturating concentrations of KCI or KBr. Kinetic analysis of these data revealed two decay processes, with time constants of τ₁≃150 μs and τ₂≃16 ms in the presence of either halide, with τ₂ describing the return to the starting (hR) state. Comparison to previous low-temperature FTIR spectra of hR intermediates confirms that characteristic hK and hL spectral features are both present before the τ₁ decay, in a state previously defined as hK_L (Dioumaev, A., and M. Braiman. 1997. Photochem. Photobiol. 66:755-763). However, the relative sizes of these features depend on which halide is present. In Br^- the hL features are clearly more dominant than in Cl^-. Therefore, the state present before τ₁ is probably best described as an hK_L/hL₁ equilibrium, instead of a single hK_L state. Different halides affect the relative amounts of hK_L and hL₁ present, i.e., Cl^- produces a much more significant back-reaction from hL₁ to hK_L than does Br^-. The halide dependence of this back-reaction could therefore explain the halide selectivity of the halorhodopsin anion pump.