Photochemistry of 124 kilodalton Avena phytochrome in vitro

The photochemical properties of purified 124 kilodalton (kD) Avena cv Garry phytochrome are examined and compared with those of the proteolytically degraded 118/114 kD species. The proportion of the chromoprotein in the far red absorbing form, Pfr, following saturating red irradiation is 0.86 for 124 kD phytochrome, substantially higher than the values of 0.79 determined here and 0.75 reported in the literature for 118/114 kD preparations. The ratio of the quantum yields for Pr to Pfr phototransformation and for Pfr to Pr phototransformation (phir/phifr) is 1.76 for the 124 kD molecule and 0.98 for the 118/114 kD species. Based on extinction coefficients determined using the Lowry assay as a measure of protein weight, the individual phototransformation quantum yields for 124 kD phytochrome are 0.17 for Pr --> Pfr (phir) and 0.10 for Pfr --> Pr (phifr). Comparison of these quantum yields with those of the 118/114 kD species (where phir = phifr = approximately 0.11) indicates that proteolytic degradation of the 124 kD molecule to the 118/114 kD species significantly affects only phir. Therefore, the lower proportion of Pfr at photoequilibrium observed for 118/114 kD preparations is explained mainly in terms of a reduced efficiency of Pr --> Pfr phototransformation. The absolute Pfr absorbance spectrum for 124 kD phytochrome obtained by correcting the measured spectrum for residual Pr exhibits a maximum at 730 nm and differs from previous absolute Pfr spectra for both ;120' kD and 60 kD phytochrome in that it lacks a shoulder in the red region of the spectrum.