ABSTRACT The crystal structure of the light-harvesting protein phycocyanin from the cyanobacterium Cyanidium caldarium with novel crystal packing has been solved at 1.65-Angstrom resolution. The structure has been refined to an R value of 18.3% with excellent backbone and side-chain stereochemical parameters. In crystals of phycocyanin used in this study, the hexamers are offset rather than aligned as in other phycocyanins that have been crystallized to date. Analysis of this crystal's unique packing leads to a proposal for phycobilisome assembly in vivo and for a more prominent role for chromophore beta-155. This new role assigned to chromophore beta-155 in phycocyanin sheds light on the numerical relationships among and function of external chromophores found in phycoerythrins and phycoerythrocyanins.
INTRODUCTION
Phycobiliproteins are bile pigment-apoprotein complexes that constitute the major light-harvesting antennae for oxygenic photosynthesis in cyanobacteria and red algae (Gantt, 1990; Glazer, 1989; MacColl and Guard-Friar, 1987). Allophycocyanin, phycocyanin, phycoerythrin, and phycoerythrocyanin, the principal phycobiliproteins, consist of aand beta-subunit polypeptides to which one or more linear tetrapyrrole chromophores are covalently attached (Bryant, 1991; Glazer, 1985; Troxler, 1994). Phycocyanobilin (PCB) is the chromophore of allophycocyanin and phycocyanin. In both proteins the chromophores are attached by cysteinyl thioether linkages at alpha-84 and beta-84 positions; in phycocyanin a second PCB is found attached to cysteine at beta-155 in a loop not found in allophycocyanin.
The alpha- and beta-subunits of phycobiliproteins exhibit a high affinity for one another and associate into alpha(beta) monomers, which in turn aggregate into (a3)3 trimers and alpha(beta)6 hexamers. In vivo, hexameric phycobiliproteins together with linker proteins self-assemble into macromolecular lightharvesting complexes (called phycobilisomes) occurring in vivo on the stroma surface of the thylakoid membranes (Bryant, 1991; Gantt, 1990; Glazer, 1989; Glazer and Melis, 1987). Phycobilisomes have two components: 1) the core, usually comprising two or three rods built from two spectroscopically distinct allophycocyanins, which are in contact with the thylakoid membrane, and 2) the rods, usually six, comprising two or more phycocyanin hexamers that radiate out from the core into the stroma space of the thylakoid membrane. In some species, rods also contain two or more hexamers of phycoerythrin or phycoerythrocyanin, which are located distally to phycocyanin hexamers. Core hexamers as well the external rods are joined together by an array of colorless linker proteins.
Phycobilisomes are very effective energy transducers (Porter et al., 1978), transferring the absorbed energy from sunlight to the photosystem II within the thylakoid membrane with more than 90% efficiency (Gantt, 1990). An understanding of the physical processes leading to such effective energy transfer would greatly enhance our knowledge of the energy transfer mechanism in photosynthesis.
Although crystals of phycobiliproteins have been available for some time (Dobler et al., 1972; Fisher et al., 1980; Hackert et al., 1977), the first structure of a phycobilisome protein was that of C-phycocyanin from Mastigocladus laminosus (Schirmer et al., 1985). Other hexameric phycobiliprotein structures solved since include C-phycocyanin from Agmenellum quadruplicatum (Schirmer et al., 1986), phycoerythrocyanin from Mastigocladus laminosus (Duerring et al., 1990), C-phycocyanin from Fremyella diplosiphon (Duerring et al., 1991), B-phycoerythrin from Porphyridium sordidum (Ficner et al., 1992), and allophycocyanin from Spirulina platensis (Brejc et al., 1995). All are remarkably similar.
Generally phycobiliprotein alpha(beta)monomers are organized into crystallographic trimers (Fig. 1), two of which come together to form a face-to-face hexamer with approximate D3 symmetry. In phycocyanin, two chromophores at positions alpha-84 and beta-84 are found toward the center of trimeric discs in regions that have structural similarity to the heme binding pocket in the globin family of proteins (Pastore and Lesk, 1990; Schirmer et al., 1985), whereas the chromophore beta-155 is located in a short loop unique to the beta-subunit of phycocyanin and is close to the edge of the trimeric disc.
Crystals of the C-phycocyanin from unicellular rhodophyte, Cyanidium caldarium, have an unusual feature: the crystal packing is different from that found in other phycocyanins. The protein molecules are arranged in layers rather than in columns. Because of the unique crystal packing, the spatial orientation of chromophores (particularly beta-155) is different, and this raises the possibility that beta-155 has a more important role in energy transfer between adjacent hexamers than commonly believed. The significance of the possible involvement of the 3-155 adduct for modes of energy transfer between the hexamers in other phycobiliproteins and for phycobilisome rods in vivo is discussed below.
MATERIALS AND METHODS
Protein purification and crystallization
Phycobilisomes from C. caldarium contain allophycocyanin (plus linker proteins) in the core and phycocyanin as the only colored protein in the rods. The phycocyanin was purified as described earlier (Offner et al., 1981; Troxler et al., 1981) and concentrated to 15 mg/ml. The purity was measured by the ratio of absorbance at 620 nm (chromophore) and 280 nm (protein) and, in protein samples used for crystallization, was more than 8. The crystals were grown by the sitting drop vapor diffusion method from 50-(mu)l droplets of IS mg/ml protein in SQ mM phosphate buffer (pH 7.0) and 0.45 M ammonium sulfate with 0.02% sodium azide equilibrated against the 1.0 M ammonium sulfate with buffer in the mother liquor. The protein crystallized into the rhombohedral R3 or R32 space group with cell dimensions of a = b = 106.42 Angstrom, c = 176.18 Angstrom, and gamma = 120 deg for the hexagonal cell. The initial precession photograph and Wilson statistics analysis were inconclusive about the space group assignment. Therefore the data were collected in the R3 symmetry.
Data collection and structure solution
A data set to 1.65-Angstrom resolution was collected with two crystals (both 0.5 x 0.5 X 0.5 mm in size) on a Hamlin area detector system (Table 1). About 78,000 unique reflections with overall R^ sub merge^ = 8.9% gave an 90.3% complete data set to 1.65-A resolution. The completeness in the last high-resolution shell of data (1.68-1.65 Angstrom) was 65%. The structure was solved by the molecular replacement method in the R3 space group. A dimer of alpha(beta)-monomers from Fremyella diplosiphon (Duerring et al., 1991 ) was retrieved from the PDB (Bernstein et al., 1977) and used as a trial structure (PDB code lcpc). The crystallographic c axis constitutes the threefold symmetry axis in each crystal. A rotational search around the crystallographic c axis with F. diplosiphon coordinates resulted in one clearly correct position with a significantly lower R factor (40.5%), which showed a rotation of 250 from the F. diplosiphon hexamer position and was chosen as a starting solution for the refinement. This position coincided with the twofold in the R32 space group. Choice of the R32 space group was confirmed by a Patterson self-rotation search (data not shown).
Refinement
The model of phycocyanin from C. caldarium was created by graphically "mutating" all nonidentical amino acids of F. diplosiphon structure, using the program CHAIN (Sack, 1988). Conformations of the "mutated" residues were adjusted to avoid interpenetrations. The refinement as well as molecular replacement were carried out using program X-PLOR 3.1. Several slow-cool cycles of refinement combined with the manual modeling after each cycle were necessary to obtain convergence. Each slow-cool cycle contained 50 cycles of Powell minimization, molecular dynamics run at 2000 K for 1 ps, and a slow cooling cycle (30 K every 50 steps), followed by 100 cycles of Powell minimization and individual temperature factor refinement. Refinement was initiated in the R3 space group, using the program X-PLOR (Brunger, 1992) as well as in the R32 space group. After three heat-cool cycles of refinement in the resolution range 10.01.65 Angstrom followed by manual rebuilding, the R factor converged to 19.5% in the R3 space group and with R = 18.3% in the R32 space group. The model refined in the R32 space group had very good stereochemistry (RMS on bonds 0.009 Angstrom and on angles 1.8o). Differences between the models were negligible; therefore, only the model refined in the R32 space group is described. The refined model has been deposited to the PDB (code lphn).
Energy transfer calculations
Theory by Forster (1967) provides the background for calculating the energy transfer rates in a weakly coupled system of chromophores by an isolated dipole approximation. Assuming the validity of the theory and neglecting the spectral differences between the chromophores, we have calculated the energy transfer rates for the C. caldarium phycocyanin crystals according to the simplified Forster formula:
RESULTS AND DISCUSSION
Description and quality of the structure
The sequence similarity between the phycocyanins from C. caldarium and from F. diplosiphon is high: 72% identity for the a-subunit and 76% identity for the beta-subunit. The changes are observed mostly on the surface of the molecules, and most of them are highly conservative (Asp-Glu, Ser-Thr, Arg-Lys). There is one exception, however. There is a patch of hydrophobic residues at the interface between the subunits comprising residues 9, 5, 24, and 27 from the alpha-subunit and residues 38, 100, and 106 from the beta-subunit. These residues are homologous but different in size between these two phycocyanins. These substitutions may be responsible for slightly different packing of alpha(beta)-subunits and the different crystal packing.
The high sequence homology suggests that the corresponding three-dimensional structures should be very similar. Indeed, the RMS difference between Cas of the starting model and of the refined structure is only 0.76 Angstrom compared to the positional errors estimated by the Luzzati method (Luzzati, 1952) of 0.2 Angstrom. This shows the strong structural homology to the F. diplosiphon phycocyanin. Nevertheless, the different crystal packing suggests that there are crucial differences between the structures. Below, we first describe the fold of the protein, then elucidate the common features of the models, and finally focus on the differences.
The chain fold for C. caldarium phycocyanin (Fig. 1) is mostly or-helical and, as indicated above, is similar to that of F. diplosiphon and other phycobiliproteins (Duerring et al., 1990; Ficner et al., 1992). Both alpha- and beta-subunits form well-defined, helical globin-like domains with six helices folded into an arrangement similar to that of myoglobin, with PCB chromophores alpha- and beta-84 (tetrapyrroles) located at sites corresponding to the heme group in myoglobin. The globin domains are complemented with the two additional helices (X, Y) (Schirmer et al., 1985), which are responsible for the formation and stability of alpha(beta)-monomers. Helices Y, A, and B can be considered as one bent helix. In the C. caldarium structure this helix has a slightly different bending angle. As a result, alpha- and beta-subunits are pinched closer to each other, and the tips of the most distant loops are closer by ~1.5 Angstrom (C(alpha)-121alpha-C(alpha)-121cr-C(alpha)-121(beta)).
In the C. caldarium phycocyanin, the globin domains are also distorted slightly and show a roll away from the threefold axis by ~ 1. 5 deg. This causes an increased ruggedness of the surface of the hexamer. The hexamer, as refined in R32 space group, has a lower and upper trimer of a-monomers related by the crystallographic twofold axis. We can speculate that the above-mentioned hydrophobic patch at the trimer-trimer interface may cause these global conformational changes.
The backbone conformation is well within allowed regions on the Ramachandran plot (Ramachandran et al., 1963), with the exception of beta-Thr77. The unusual conformation of this residue, which is located on the loop enveloping chromophore beta-84, is caused by the close contact with the chromophore alpha-84 of the neighboring monomer. The backbone conformation of this residue in the C. caldarium phycocyanin structure (phi = 86.7 deg , psi = 153.3 deg) agrees well with previous findings (Duerring et al., 1991).
Methylation of beta-Asn72 was also detected as before (Duerring et al., 1991; Klotz et al., 1986). Asn72 was initially sequenced as Thr (Offner et al., 1981); later this misassignment was corrected when the DNA sequence became available (RFT, unpublished data). The side chain of Asn72 provides a direct link of the chromophore beta-84 to solvent. It has been proposed that this conserved methylation serves a functional role (Swanson and Glazer, 1990) by providing a better separation of the chromophore from the solvent. One can easily speculate that the electronic properties of the beta-84 chromophore would have been different without the methylation, which consequently would lead to a less efficient chromophore.
The electron density is of sufficiently high quality to locate every nonhydrogen atom (Figs. 2 and 3). This high quality electron density allowed us to model more than 500 water molecules, some deep in the protein interior, using the difference electron density maps. The criterion adopted was that the water molecule was incorporated into the model if the electron difference density was above 2.4sigma. Electron density is weak and disconnected only for residues 115-123 in the beta-subunit. This sequence contains the single free cysteine in the structure. The backbone temperature factors in this region were above 40 Angstrom2, whereas elsewhere they were below 20 Angstrom2.
Chromophores
Crystal contacts
Although F. diplosiphon and C. caldarium crystallized in similar rhombohedral space groups (R3 and R32, respectively), there is a profound difference in the lattice dimensions and cell organization. In crystals of phycocyanin from C. caldarium the hexamers are offset, whereas in F. diplosiphon crystals they are organized in columns with a crystallographic translational repeat of 61.24 Angstrom along the c axis. The hexamers in C. caldarium crystals have unique stacking contacts along the body diagonal. This offset stacking along the cell diagonal allows the hexamers to pack over one another more tightly than in F. diplosiphon because the hexamers are effectively rotated relative to one another. The tighter packing is apparent from the smaller average hexamer-hexamer distance along the c axis in C. caldarium crystals of 58.72 Angstrom (176.18/3), as compared to 61.24 Angstrom for F. diplosiphon. The tight packing of hexamers in C. caldarium phycocyanin crystals results in crystals that are very stable mechanically, which diffract well, even when heated above 50C.
Modeling hexamers in the phycobilisome rods
Because in other phycocyanins that crystallized in rhombohedral space groups the hexamers are organized in columns due to the crystal translational repeat, it has been suggested (e.g., Schirmer et al., 1986) that the organization of hexamers in vivo is similar to what occurs in crystals. The exact translational repeat precludes the possibility that hexamers in columns could be rotated by a small angle (Fig. 5) to adjust the contacts. However, efficient packing of hexamers would be required in the native antenna to keep the exposed phycobilisome rods stable in the surrounding solvent.
To determine the optimal van der Waals packing of phycocyanin hexamers in columns (rods), a hexamer contact search was performed by rotating one hexamer on top of another in one degree steps over a 120 deg range, starting with exactly aligned hexamers as in the F. diplosiphon. The contacts were optimized at every step by shifting the top hexamer up and down so that the closest approach of the Ca in rotated hexamers would be within 5-5.5 Angstrom. The result of this optimization is presented in Fig. 5 A. At the minimum, which was located ~30 deg from the initial position, the hexamers were separated by 56.18 Angstrom.
The hexamers in C. caldarium have slightly more rugged surfaces than those of the phycocyanin from F. diplosiphon. This ruggedness is associated with the alpha(beta) packing differences (see above). To evaluate the contribution of electrostatic energy to the stability of the rods composed of optimally packed, rotated hexamers, we calculated the electrostatic field and projected it onto the molecular surface (Fig. 6). The regions in contact in the rotated adjacent hexamers are marked by green bars. Visual inspection shows that these red (negative) and blue (positive) charged patches at the knobs and holes, respectively, on the hexamer surface would interact favorably (Fig. 6). This observation agrees also with the tentative conclusions of the electrostatic calculations carried out on the F. diplosiphon protein (Karshikov et al., 1991). Karshikov et al. find a shallow minimum. Although their global minimum in hexamerhexamer rotation is 0 deg , the 30 deg rotated position is less than 1 kcal higher in energy. Note that for phycocyanin the dominant electrostatic field is negative (red), particularly in the center of the hexamers. This charge distribution may be important for efficient energy transfer and for strong binding of the linker proteins (Lundell et al., 1981).
We propose that the same structural feature of rotated hexamers that is used in the crystal lattice of C. caldarium phycocyanin is important for the native phycobilisome rods. The difference between the arrangement of hexamers packed into rods proposed by others (e.g., Schirmer et al., 1986) and that proposed here can clearly be seen in Fig. 7. Considerably more space is found between the hexamers in Fig. 7 A, where the hexamers are not rotated, as compared to Fig 7 B, where rotation to the optimal angle of 30 deg is used. The tighter packing of the rotated rods would be stabilized not only by van der Waals contacts, but by electrostatic interactions, as just noted. Furthermore, this rotated hexamer packing has implications for the importance of each chromophore in energy transfer along the rods.
Energy pathways
In phycobilisomes, energy is transferred along the rods to the core by the chromophores. Chromophores are divided into two spectroscopic classes (Teale and Dale, 1970). The s (sensitizing) class absorbs at the blue edge of the absorption band and transfers the excitation energy in a radiationless (resonant) manner to the f (fluorescing) chromophores. Excitation at the red absorption edge by the f-chromophores results in depolarization, which suggests that the energy is transferred in the rods along the line of f-chromophores.
Using Forster theory (Forster, 1967), Schirmer et al. (1986), as well as others (Sauer and Scheer, 1988), have calculated the energy transfer rates for the different systems of chromophores in different crystals and concluded that chromophore 1-84 is a type f chromophore. Analysis of the chromophore distances in C. caldarium lattice suggests that chromophore 3-155 should play a more significant role than that ascribed to it before.
The calculations discussed here are not intended to compete with more sophisticated methods such as those presented by Sauer and Scheer, (1988) but were meant to provide the background for the structural as well as functional comparisons. The levels of these calculations are nevertheless fully compatible with those presented by Schirmer et al. (1986) and Duerring et al. (1990, 1991). In fact, the orientation factors and transmission rates are very similar in all of those species, despite small structural differences. The values presented in Table 2 should be compared with those presented in figure 11 and table 9 of Duerring et al. (1991).
In Fig. 8, we compare the chromophore packing schemes in the crystal lattices of F. diplosiphon and C. caldarium. Table 2 gives the distances, orientation factors, and transition rates calculated from the Forster theory for the C. caldarium lattice. The beta-84 chromophore distances are similar in the two lattices. A comparison indicates that the only significant difference is the closeness of chromophores alpha-84 and beta-155 between translationally related molecules in the lattice (26.7 Angstrom, 30.4 Angstrom) (Fig. 8). These contacts are absent in the F. diplosiphon lattice.
Recent results have suggested that chromophores alpha-84 and beta-84 are the ones most involved in interhexamer energy transfer and that beta-84 is of type f (Schirmer et al., 1987; Siebzehrubl et al., 1987). In fact, the distance between these chromophores (~20 Angstrom) would indicate that the excitonic coupling should occur, and both chromophores should be electronically coupled. However, those results were obtained on isolated dimers and trimers and do not put strict constrains on the hexamer interactions. In the optimally packed hexamers chromophores beta-84 of the neighboring hexamers are properly oriented and at appropriate distances (~30 Angstrom), so they can efficiently exchange energy by the FOrster mechanism. Energy transfer rates calculated for chromophores beta-155 in optimally packed hexamers (Fig. 7) have interhexamer energy transfer along the beta-155 chromophore path only slightly less efficient than along the line of the beta-84-alpha-84 path (Table 3). This would create an additional pathway on the periphery of the hexamers, which would involve the chromophore beta-155.
The proposed role of chromophore beta-155 would lead to a modified picture of an auxiliary energy transfer pathway along the outer surface of phycobilisome rods (Fig. 9) in addition to the dominant one along the linker proteins filling the inner space of the rods (Glazer, 1989; MacColl and Guard-Friar, 1987). However, because this auxiliary pathway is more exposed to solvent, it must be less efficient than the internal fully isolated pathway along chromophores beta-84. Nevertheless, this proposal would offer a tentative answer to the question of why additional external chromophores are found more frequently in phycobilisomal proteins located closer to the tip of the rods (phycoerythrins and phycoerythrocyanins). Such external chromophores would collect light efficiently and then partially transfer the energy to the core and partially conduct it along the surface of the rods through an array of other external chromophores. The spectroscopic ladder formed by the different types of chromophores with descending order of absorbed and emitted wavelengths would promote an efficient transfer to the core on both internal and external pathways. Decreasing the number of external chromophores while forming an extensive antenna system would also form an "energy funnel" that would eventually lead to an efficient energy transfer to the core.
In assessing the importance of the new pathways for energy transfer at physiological conditions, the role of the linker proteins on the energy transfer capabilities of isolated hexamers versus intact rods must also be considered. Relatively little is known about the structural features of the linker proteins, except for the fact that they do not have chromophores and they have to modify the structure of colored proteins to achieve a high quantum efficiency of energy transfer not seen in isolated dimers, trimers, or hexamers. To address this question we had purified phycocyanin with at least one linker protein (27 kDa), as testified by absorption spectroscopy and by the sodium dodecyl sulfate gel chromatography of the initial material and the dissolved crystals, and crystallized it. Unfortunately this 27-kDa linker protein did not influence the crystal packing arrangement, and the crystals showed R3 (R32) space group. Therefore, it meant that the linker protein did not have any external contacts and was three-way disordered in the crystal and hence invisible. In those crystals the positions of chromophores were identical to the pure alpha(beta) dimer crystal structure, which argued that these particular linker proteins had no influence on the geometric characteristics of the hexamers. The spectrum of the complex (red-shifted with a shorter wavelength shoulder, as observed before; Lundell et al., 1981) did not change much in the isolated hexamers. This would argue that the transmission pathways in isolated hexamers are not greatly influenced much by linker proteins. Therefore, we would expect that a complete three-dimensional assembly of the phycobilisomal rod has a much greater influence on energy transfer and spectroscopic characteristics.
The hypothesized auxiliary pathways and the importance of external chromophores could be tested experimentally by mutagenesis of beta-155 cysteine, the external chromophore attachment site, and spectroscopic measurement of the energy transfer through the reconstituted rods. Such an experiment would answer an important question about energy transduction by phycobilisome rods. The first step had already been taken by Debreczeny et al. (1993), who measured absorption and emission spectra of individual chromophores in alpha(beta)-monomers of the wild type and in the Cysl55-to-Ser mutant of cyanobacterium Synechocus sp.
This work was supported by grant R01-GM38114 to MMT from the National Institutes of Health. BS acknowledges support from this source. This work was also supported by grant DCB9119062 to RFT from the National Science Foundation. This work was also supported in part by a training fellowship from the Keck Center for Computational Biology to BS (National Science Foundation RTG grant BIR-94-13229).
[Reference]
REFERENCES
[Reference]
Bernstein, F. C., T. F. Koetzle, G. J. B. Williams, E. F. Meyer, Jr., M. D. Brice, J. R. Rodgers, O. Kennard, T. Shimanouchi, and M. Tasumi. 1977. A computer based archival file for macromolecular structures. J. Mol. Biol. 112:535-542.
Brejc, K., R. Ficner, R. Huber, and S. Steinbacher. 1995. Isolation, crystallization, crystal structure analysis and refinement of allophycocyanin from the cyanobacterium Spirulina platensis at 2.3 A resolution. J. Mol. Biol. 249:424-440.
[Reference]
Brunger, A. T. 1992. X-Plor Version 3.0: A System for Crystallography and NMR. Yale University, New Haven, CT.
Bryant, D. A. 1991. Cyanobacterial phycobilisomes: progress towards a complete structural and functional analysis via molecular genetics. In The Photosynthetic Apparatus, Molecular Biology and Operation, Cell Culture and Somatic Cell Genetics of Plants, Vol. 7B. L. Bogorad and I. K. Vasil, editors. Academic Press, New York. 255-298. Debreczeny, M., K. Sauer, J. Zhou, and D. A. Bryant. 1993. Monomeric C-phycocyanin at room temperature and 77K: resolution of the absorption and fluorescence spectra of individual chromophores and energytransfer rate constants. J. Chem. Phys. 97:9852-9862.
[Reference]
Dobler, M., S. D. Dover, K. Laves, A. Binder, and H. Zuber. 1972. Crystallization and preliminary crystal data of C-phycocyanin. J. Mol. Biol. 71:785-787.
Duerng, M., R. Huber, B. Wolfram, R. Ruembeli, and H. Zuber. 1990. Refined three-dimensional structure of phycoerythrocyanin from the cyanobacterium Mastigocladus laminosus at 2.7 A. J. Mol. Biol. 211: 633-644.
[Reference]
Duerring, M., G. B. Schmidt, and R. Huber. 1991. Isolation, crystallization, crystal structure analysis and refinement of constitutive C-phycocyanin from the chromatically adapting cyanobacterium Fremyella diplosiphon at 1.66 A resolution. J. Mol. Biol. 217:557-591. Ficner, R., K. Lobeck, G. Schmidt, and R. Huber. 1992. Isolation, crystallization, crystal structure analysis and refinement of B-phycoerythrin from the red alga Porphyridium sordidum at 2.2 A resolution. J. Mol. Biol. 228:935-950.
[Reference]
Fisher, R. G., N. E. Woods, H. E. Fuchs, and R. M. Sweet. 1980. Three-dimensional structures of C-phycocyanin and B-phycoerithrin at 5-ti resolution. J. Biol. Chem. 255:5082-5089. Forster, T. 1967. Mechanisms of energy transfer. In Comprehensive Biochemistry, Vol. 22. M. Florkin and E. H. Stotz, editors. Elsevier, Amsterdam. 61-80.
Gantt, E. 1990. Pigmentation and photoacclimation. In Biology of Red Algae. K. M. Cole and R. G. Sheath, editors. Cambridge University Press, New York. 203-219.
[Reference]
Glazer, A. N. 1985. Light harvesting by phycobilisomes. Annu. Rev. Biophys. Chem. 14:44-77.
Glazer, A. N. 1989. Light guides. Directional energy transfer in a photosynthetic antenna. J. BioL Chem. 264:1-4.
Glazer, A. N., and Melis, A. 1987. Photochemical reaction centers; structure, organization and function. Annu. Rev. Plant PhysioL 38:11-45. Hackett, M. L., C. Abad-Zapatero, S. J. Stevens, and J. L. Fox. 1977. Crystallization of C-phycocyanin from the marine blue-green alga Agmenellum quadruplicatum. J. MoL BioL 111:365-369.
[Reference]
Karshikov, A., M. Duerring, and R. Huber. 1991. Role of electrostatic interaction in the stability of the hexamer of constitutive phycocyanin from Fremyella diplosiphon. Protein Eng. 4:681-690. Klotz, A., J. A. Leary, and A. Glazer. 1986. Post-translational methylation of asparaginyl residues. Identification of 3-71 y-N-methylasparagine in allophycocyanin. J. Biol. Chem. 261:15891-15894. Lundell, D. J., R. C. Williams, and A. N. Glazer. 1981. Molecular architecture of light-harvesting antenna. J. BioL Chem. 256:3580-3592. Luzzati, V. 1952. Traitement statistique des erreurs dans la determination des structures cristallines. Acta Crystallogr. 5:802-810. MacColl, R., and D. Guard-Friar. 1987. Phycobiliproteins. CRC Press, Boca Raton, FL.
[Reference]
Offner, G. D., A. S. Brown-Mason, M. M. Ehrhardt, and R. F. Troxler. 1981. Primary sequence of phycocyanin from the unicellular Rhodophyte Cyanidium caldarium. I. Complete amino acid sequence of the a subunit. J. Biol. Chem. 256:12167-12175.
Pastore, A., and A. M. Lesk. 1990. Comparison of the structures of globins and phycocyanins: evidence for evolutionary relationship. Proteins Struct. Funct. Genet. 8:133-155.
Porter, G., C. J. Tredwell, G. F. Searle, and J. Barber. 1978. Picosecond time-resolved energy transfer in Porphiridium cruentum. Part I In intact alga. Biochim. Biophys. Acta. 501:232-245. Ramachandran, G. N., C. Ramakrishnan, and V. Sasisekharan. 1963. Stereochemistry of polypeptide chain configurations. J. MoL BioL 7:95-99.
[Reference]
Sack, J. S. 1988. Chain-a crystallographic modeling program. J. Mol. Graph. 6:244-245.
Sauer, K., and H. Scheer. 1988. Excitation transfer in C-phycocyanin. Forster transfer rate and excitation calculations based on a new crystal structure data for C-phycocyanin from Agmenellum quadruplicatum and Mastigocladus laminosus. Biochim. Biophys. Acta. 936:157-170. Schirmer, T., W. Bode, and R. Huber. 1987. Refined three-dimensional structures of two cyanobacterial C-phycocyanins at 2.1 and 2.5 tL resolution. J. Mol. Biol. 196:677-695.
[Reference]
Schirmer, T. D., W. Bode, R. Huber, W. Sidler, and H. Zuber. 1985. X-ray crystallographic structure of the light-harvesting biliprotein Cphycocyanin from thermophilic cyanobacterium Mastigocladus laminosus and its resemblance to globin structures. J. Mol. Biol. 184:257-277. Schirmer, T., R. Huber, M. Schneider, W. Bode, M. Miller, and M. L. Hackett. 1986. Crystal structure analysis and refinement at 2.5 A of hexameric C-phycocyanin from the cyanobacterium Agmenellum quadruplicatum. The molecular model and its implications for lightharvesting. J. Mol. Biol. 188:651-676.
Siebzehrubl, S., R. Fisher, and H. Scheer. 1987. Chromophore assignment in C-phycocyanin from Mastigocladus laminosus. Z. Naturforsch. 42C: 258-262.
[Reference]
Swanson, R. V., and A. N. Glazer. 1990. Phycobiliprotein methylation. Effect of the gamma-N-methylasparagine residue on energy transfer in phycocyanin and phycobilisome. J. Mol. Biol. 214:787-796. Teale, F. W. J., and R. E. Dale. 1970. Isolation and spectral characterization of phycobiliproterins. Biochem. J. 116:161-169. Troxler, R. F. 1994. Molecular aspects of pigments and photosynthesis in C. caldarium (Rhodophyte) and related cells. In Evolutionary Pathways and Enigmatic Algae: Cyanidium caldarium (Rhodophyta) and related cells. J. Seckback, editor. Kluwer Academic Publishers, Dordrecht, the Netherlands.
Troxler, R. F., M. M. Ehrhardt, A. S. Brown-Mason, and G. D. Offner. 1981. Primary sequence of phycocyanin from the unicellular Rhodophyte Cyanidium caldarium. II. Complete amino acid sequence of the 3 subunit. J. Biol. Chem. 256:12176-12184.
[Author Affiliation]
Boguslaw Stec,* Robert F. Troxler,# and Martha M. Teeter*
*Department of Chemistry, Merkert Chemistry, Merkert Chemistry Building, Boston College, Chestnut Hill 02167 and #Department of Biochemistry, Boston University, School of Medicine, Boston, Massachusetts 02118 USA
[Author Affiliation]
Received for publication 7 July 1998 and in final form 23 February 1999. Address reprint requests to Dr. Boguslaw Stec, Department of Chemistry and Cell Biology, Rice University, Mail Stop 140, 6100 Main St., Houston, TX 77005. Tel.: 713-527-8101, ext. 3346; Fax: 713-285-5154; E-mail: stec@bioc.rice.edu.
1999 by the Biophysical Society 0006-3495/99//06/2912/10 $2.00
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