B850 pigment-protein complex of Rhodopseudomonas sphaeroides: Extinction coefficients, circular dichroism, and the reversible binding of bacteriochlorophyll

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Chromatophores of Rhodopseudomonas sphaeroides yield the antenna complex B850 in either of two states, depending on the method of isolation. Methods using dodecyl (= lauryl) dimethylamine oxide yield B850 with an absorption spectrum like that in vivo: the bands at 800 and 850 nm, due to the bacteriochlorophyll (Bchl) components Bchl-800 and Bchl-850, are in ratio A800/A850 = 0.65 ± 0.05. When B850 is isolated by methods using dodecyl sulfate, the Bchl-800 is attenuated or absent. Bchl assays of these materials and of the isolated antenna complex B875 yielded the following extinction coefficients, ±SD, on the basis of the molarity of Bchl: For B875, ε875 = 126 ± 8 mM-1 cm-1. For B850 in the normal (high-Bchl-800) state, ε850 = 132 ± 10 mM-1 cm-1. For the individual components of Bchl in B850, ε850 of Bchl-850 = 184 ± 13 mM-1 cm-1 and ε800 of Bchl-800 = 213 ± 28 mM-1 cm-1. With these coefficients the molecular ratio of Bchl-850 to Bchl-800 equals 1.8 ± 0.4 for B850 in the high-Bchl-800 state. Starting with B850 depleted of Bchl-800, the addition of dodecyldimethylamine oxide restored the 800-nm absorption band. The 850-nm band became shifted toward the blue, narrowed, and slightly attenuated, and its associated circular dichroism became 20% more intense. Free Bchl added with dodecyldimethylamine oxide accelerated the restoration of Bchl-800 and retarded the attenuation of Bchl-850. We conclude that free Bchl can interact reversibly with a binding site for Bchl-800 in the B850 complex, with dodecyl sulfate favoring dissociation and dodecyldimethylamine oxide promoting association. Thus the reversible dissociation of a native chlorophyll-protein complex has now been demonstrated.

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