Binding of phytochrome to plasma membranes in vitro

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Abstract
Plasma membranes were prepared from dark grown shoots of oat (Avena sativa, L. cv. Sol II) by means of an aqueous polymer two‐phase partition method. Phytochrome was prepared from the same material by brushite chromatography and immunoaffinity chromatography. The purified plasma membranes and the phytochrome were mixed and pelleted by centrifugation at 72 000 g. The pellet was washed and repelleted once, and the phytochrome found in the resulting pellet was regarded as specifically bound to the plasma membrane. This binding was increased in the presenee of Mg2+ and after red irradiation. The affinity constant was 3.4 ± 106M‐1 as extrapolated from a Scatchard plot. An increased binding in the presence of 1 M sodium chloride indicates that the binding is of a hydrophobic nature. A tryptic treatment increased the binding which could be interpreted as the removal of steric hindrance from the membranes and as the non‐existence of a trypsin‐degradable receptor protein.