ISOLATION AND CHARACTERIZATION OF THE CORE LIGHT-HARVESTING COMPLEX B875 AND ITS SUBUNIT FORM, B820, FROM RHODOCYCLUS-GELATINOSUS

We present a new method of preparation for Rhodocyclus gelatinosus lig ht-harvesting complex B875 and its subunit form B820, without prior ex traction of carotenoids from chromatophores. This method was based on partial dissociation of isolated B875 into B820 during hydrophobic chr omatography in presence of ammonium sulphate. The two forms were then separated by gel chromatography in presence of this salt, which was re quired for stabilising B820. Carotenoid composition was determined for each form. B875 contained two hydroxyspheroidene molecules per bacter iochlorophyll dimer. Carotenoid content of B820 was about 5-times lowe r, and variable, indicating that most of this pigment was lost during the formation of B820. The apparent molecular mass of B820 antenna, de termined by gel filtration, was 18-25 kDa. For B875 several values wer e obtained, depending on chromatographic conditions (protein and ammon ium sulphate concentrations): > 600 kDa, 130 kDa and 35-40 kDa. These values may be inaccurate, because of the unknown contribution of deter gent and possible interactions between the protein and the gel matrix. The results, however, indicated that B875 could present several state s of aggregation, and that B820 has a smaller apparent molecular mass. Absorbance and CD spectra were measured and compared with spectra pub lished for other species. We did not observe reconstitution of the ori ginal B875 form by increasing B820 concentration or decreasing ammoniu m sulphate concentration. However, another spectral form with absorban ce maximum between 830-845 nm was observed, with a CD spectrum differe nt in the infrared region from that of B820. A possible requirement of carotenoid for reassociation of B820 into B875 is discussed.