Proteolytic modifications of the B800-860 complex of the photosynthetic bacterium Rhodocyclus tenuis: Structural and spectral effects

Abstract

The modification effects on the absorption and cirular dichroic (CD) spectra of the isolated B800-860 antenna complex of Rhodocyclus tenuis by a number of proteolytic enzymes were investigated. The chymotrypsin modifications of the B800-860 complex led to an about 40% decrease of the 860-nm band and a blue-shift to 841 nm. The biphasic CD signal related to the B860 BChl disappeared and a new double CD signal with a zero-crossing point at 842 nm appeared. These absorption and CD spectral changes suggested that a B800-841 complex resulted after chymotrypsin digestion. The polypeptide components of the chymotrypsin-modified B800-860 complex were separated by reverse-phase chromatography, and their amino acid sequences determined by protein sequencing and mass spectrometry. Sequence analyses showed that the C-terminal 25 residues of the B800-860-α polypeptide and the C-terminal 8 residues of the B800-860-β polypeptide were cleaved by chymotrypsin, and the remaining α, β polypeptide fragments apparently form the structural basis for the newly-formed B800-841 complex. No significant spectral change was observed from exposing the isolated B800-860 complex to trypsin, carboxypeptidase A and the combination of carboxypeptidase A and carboxypeptidase B. Short-term proteinase K incubation of the B800-860 complex of Rc. tenuis led to a preferential decrease of the 860-nm absorbance band and its related CD signals, as compared to the 800-nm absorbance and CD bands, suggesting that the C-terminal portions of the antenna polypeptides are possibly exposed to the exterior of the B800-860 complex micelles. Whereas, long-term proteinase K digestion resulted in the spectral collapse of the B800-860 complex and the release of free BChls. Our proteolysis experiments support the hypothesis that the C-terminal portions of the antenna polypeptides play a key role in the redshift and strong molar extinction of the Q_y band of the B850 BChls.