Some Properties of Phytochrome Isolated From Dark-grown Oat Seedlings (Avena sativa L.)

Phytochrome was partially purified from etiolated seedlings of Avena sativa L. Several properties of the red-absorbing (P(R)) and far-red absorbing (P(FR)) forms of the pigment were compared. The 2 forms could not be shown to differ with respect to their sedimentation velocity in sucrose density gradients, elution volume from Sephadex G-200 columns, binding properties on calcium phosphate, or electrophoretic mobility. P(FR), however, was more labile than P(R) during precipitation with 50% ammonium sulfate. Sephadex G-200 elution diagrams obtained with fresh phytochrome preparations revealed 2 components of different molecular weights, 1 roughly 180,000, and 1 roughly 80,000. Native phytochrome had an absorption spectrum in vivo showing an absorption maximum for P(R) of 667 nm. Both the large and small forms of phytochrome mentioned above can be maintained with an absorption maximum for P(R) of 667 nm. However, allowing them to remain for several hours as P(FR), even at 4 degrees , shifted this peak to 660 nm. The protein conformational change during phytochrome transformation may be quite small, though the various comparative techniques used do not strictly rule out a fairly large one. The need for maintaining the pigment as P(R) during all steps of purification, but particularly during ammonium sulfate precipitation is underscored.