EVIDENCE FOR A SEQUENTIAL PATHWAY FROM Pr TO Pfr OF THE PHOTOTRANSFORMATION OF 124‐kDa OAT PHYTOCHROME

Phototransformation kinetics of 124‐kDa oat phytochrome at 298 K after a red (660‐nm) laser flash excitation were recorded at different wavelengths. The kinetics of the dark relaxation processes for lumi‐R to Pfr can be satisfactorily described by only 3 rate constants: k= 28000 s‐1 370 s‐1 and 20 s‐1. The first rate constant is due to the decay of lumi‐R to meta‐Ra. The latter two rate constants correspond to processes establishing the far‐red (>700 nm) absorption band. No meta ‐Rb could be detected. From the wavelength dependency of the amplitudes of these two rates, parallel pathways in the formation of Pfr could be excluded. A unique sequential pathway for the dark relaxation leading to Pfr seems to be an intrinsic property of 124‐kDa phytochrome, however. Assuming a sequential pathway, molar extinction coefficients for intermediates have been calculated. These values agree with molar extinction coefficients obtained from low‐temperature spectra. The process with a rate constant of 370 s‐1 corresponds to absorbance changes for the formation of meta‐Rc from meta‐Ra and the rate constant of 20 s‐1 describes the absorbance changes due to the transformation of meta‐Rc to Pfr.