Isolation of a carotenoid-containing sub-membrane particle from the chloroplastic envelope outer membrane of pea (Pisum sativum).
Chloroplastic envelope membranes isolated from pea (Pisum sativum) leaves are rich in carotenoids, containing approximately 2 micrograms of carotenoid mg-1 protein. We report here that envelopes can be surfactant-solubilized while maintaining association of carotenoids with protein components of the membrane. Treatment of isolated chloroplastic envelope membranes with 0.5% Deriphat 160 (N-lauryl-beta-imminodipropionate) causes general solubilization but preserves an envelope sub-membrane fragment which is fractionated by centrifugation in a sucrose gradient and by chromatography on a column of DEAE-Sephacel. The isolated submembrane complex contained five major proteins with M(r) values equivalent to 75,000, 36,000, 34,000 17,500, and 14,500. Spectroscopic and chromatographic analyses revealed that the complex contains violaxanthin and at least one other carotenoid. Carotenoid content of the fractionated complex was estimated as 4.8 micrograms mg-1 protein. Immunoblot analysis reveals that the constituent proteins of this complex are derived from the chloroplastic outer envelope membrane. These data suggest that at least some of the carotenoids of the chloroplastic envelope may be organized by apoproteins.
Markwell
J
J
Department of Botany, University of Wisconsin, Madison 53706.
Bruce
B D
BD
Keegstra
K
K
D002338:Carotenoids / Q000302:isolation & purification*; D002458:Cell Fractionation; D002499:Centrifugation, Density Gradient; D002736:Chloroplasts / Q000737:chemistry* / Q000648:ultrastructure; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D007887:Fabaceae / Q000737:chemistry*; D007425:Intracellular Membranes / Q000737:chemistry* / Q000648:ultrastructure; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification; D008970:Molecular Weight; D010946:Plants, Medicinal; D011485:Protein Binding
D008565:Membrane Proteins; D002338:Carotenoids