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This study was designed to yield data on the supramolecular organization of the phycobilisome apparatus from Synechocystis, and the possible effects of environmental stress on this arrangement. Phycobilisomes were dissociated in a low ionic strength solution and a quantitative estimation of the protein components present in each subcomplex was obtained using liquid chromatography coupled on-line with a mass spectrometer equipped with an electrospray ion source (ESI-MS). An advantage of this approach is that information can be collected on the initial events, which take place as this organism adapts to environmental changes. Ultracentrifugation of whole phycobilisomes revealed five subcomplexes; the lightest contained four linker proteins plus free phycocyanin, the second the core complex, while the last three bands contained the rod complexes. Four linkers were found in band 1 with higher molecular masses than those expected from the DNA sequence, indicating that they also contain linked chemical groups. UV-B irradiation specifically destroyed the b-phycocyanin and one rod linker, which resulted in the disintegration of the rod complexes. The two bilins present in b-phycocyanin give a greater contribution to the UV absorption than the single bilin of the other bilinproteins and probably react with atmospheric oxygen forming toxic radicals. The protein backbone is, in fact, protected from damage in anaerobic conditions and in the presence of radical scavengers. Cells grown in sulfur-and nitrogen-deficient medium contained significantly reduced levels of b-phycocyanin and one rod linker.

. radical but also of radicals generated on the donor side. Difference analyses indicated that one of these radicals is likely to be due to a D1 tyrosine 161 or D2 tyrosine 161. In contrast, the monomeric CP47-RC complex did not show similar EPR-detectable radicals and instead was dominated by a high yield of the spin-polarized triplet signal generated by recombination reactions between the oxidized primary reductant, pheophytin, and the primary donor, P680. It is also concluded from EPR analyses that both the monomeric and dimeric forms of the CP47-RC subcore complex contain one cytochrome b 559 per reaction center.Overall the results suggest that photosystem II normally functions as a dimer complex and that monomerization at the level of the CP47-RC subcore complex leads to destabilization of the bound plastoquinone, which functions as Q A .More than 25 different protein subunits make up the photosystem II (PSII) 1 complex of oxygenic photosynthetic organisms (1). At the heart of this complex is the reaction center (RC) consisting of the D1 and D2 proteins, where primary charge separation occurs (2). Closely associated with the D1 and D2 proteins are two similar chlorophyll a-binding proteins, CP43 and CP47 (3). These proteins serve as an "inner antennae" system that is linked to a secondary light-harvesting system. In higher plants and green algae, the chlorophyll a/bbinding proteins (encoded by nuclear located cab genes) act as the secondary light-harvesting system, while phycobilisomes serve the same purpose in other types of oxygenic photosynthetic organisms, such as red algae and cyanobacteria (4). CP43 and CP47 are also distinguished by having a large hydrophilic loop linking putative membrane-spanning regions 5 and 6 (3). These loops are almost certainly located on the luminal surface of the complex and may function in water splitting in some way (5). Treatments with detergents can peel away the various subunits, and it has been shown that during such manipulations CP43 is more readily removed than CP47 (6, 7). It is therefore possible to isolate a CP47-RC complex. Recently a method was described for spinach that yielded a preparation of the CP47-RC complex consisting of a mixture of monomeric and dimeric forms (8). Analyses using mass spectrometry showed that both forms of this subcore complex contained the products of the psbE, psbF, psbI, psbT c , and psbW genes as well as the D1 and D2 proteins and CP47. However, the CP47-RC dimer contained, in addition, the products of the psbL and psbK genes. Also of significance was the finding that the dimer and not the monomer contained about one molecule of plastoquinone-9 per RC. Overall, the findings suggest that the monomeric form of the isolated CP47-RC complex was derived by dissociation of the dimer and that the latter conformation is likely to be the in vivo state. Moreover, the finding that the CP47-RC complexes contain several small proteins with putative one transmembrane helices is relevant to de-

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The electrophoretic migration behavior of three closely related hydrophobic intrinsic membrane proteins of the photosystem II light-harvesting complex (LHC II) was investigated in free solution capillary electrophoresis at pH 8.0-10 with running electrolyte solutions containing either anionic, zwitter-ionic or nonionic detergents. The complete and repeatable separation of these proteins was accomplished with a running electrolyte solution of 25 mM Tris/192 mM glycine, pH 8.8, containing either sodium dodecyl sulfate or n-octyl beta-D-glucopyranoside at concentration up to 5.0 and 7.0 mM, respectively. Migration times and resolution of the individual LHC II intrinsic membrane proteins were sensitive to the type of detergent. The effect of detergent concentration on the electrophoretic behavior of the LHC II proteins was also investigated. Electroelution of the LHC II components separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis was used to isolate these intrinsic membrane proteins, which were then injected onto the capillary electrophoresis system for peak identification.