THE WHITE MEMBRANE OF CRYSTALLINE BACTERIOOPSIN IN HALOBACTERIUM HALOBIUM STRAIN R1mW AND ITS CONVERSION INTO PURPLE MEMBRANE BY EXOGENOUS RETINAL

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Abstract
Abstract— A new strain isolated from Halobacterium halobium designated R1mW, contained negligible amounts of isoprenoid pigments, had a yellowish white color due to respiratory pigments and showed no proton movement in response to light. However, addition of all‐trans‐retinal converted R1mW into purple cells. Formation of both halorhodopsin and bacteriorhodopsin was indicated by induction of light‐dependent proton uptake and release, respectively. Both haloopsin and bacterioopsin were thus postulated to be present in R1mW. Electron micrographs of freeze‐fractured cytoplasmic membranes revealed patches in a hexagonal array of trimeric particles, comparable to the purple membrane structure.These white membrane patches were isolated by procedures similar to those for the purple membrane. The white membrane's buoyant density was about 1.18 g/m/, and its main component migrated on sodium dodecylsulfate polyacrylamide gels at the same rate as bacteriorhodopsin. The white membrane showed only a small absorption peak at ∼410nm due to contaminating respiratory pigments and a strong absorption at around 275 nm and shorter wavelengths. The white membrane was thus considered to be mainly composed of bacterioopsin, which was readily converted into bacteriorhodopsin by an addition of all‐trans‐retinal.The absorption and CD spectra of the white membrane were measured before and after addition of retinal. The molar extinction coefficient of dark‐adapted bacteriorhodopsin formed was determined to be 53000M−1 cm−1 at 560 nm from retinal binding studies. The CD spectrum of the white membrane was negligible in the visible region but showed several bands assigned to aromatic and backbone structures in the UV region. Retinal addition caused considerable changes in the spectrum, yielding the CD spectrum of crystalline purple membrane bacteriorhodopsin. The white membrane thus appears to be a preparation suitable for structure‐function studies of bacteriorhodopsin.