Herbert Zuber

X-ray crystallographic structure of the light-harvesting biliprotein C-phycocyanin from the thermophilic cyanobacterium Mastigocladus laminosus and its resemblance to globin structures

The structure of the biliprotein C-phycocyanin from the thermophilic cyanobacterium Mastigocladus laminosus has been determined at 3 A resolution by X-ray diffraction methods. Phases have been obtained by the multiple isomorphous replacement method. The electron density map could be improved by solvent flattening and has been interpreted in terms of the amino acid sequence. The protein consists of three identical (alpha-beta)-units which are arranged around a threefold symmetry axis to form a disc of approximate dimensions 110 A X 30 A with a central channel of 35 A in diameter. This aggregation form is supposed to be the same as that found in the rods of native phycobilisomes. Both subunits, alpha and beta, exhibit a similar structure and are related by a local twofold rotational axis. Each subunit is folded into eight helices and irregular loops. Six helices are arranged to form a globular part, whereas two helices stick out and mediate extensive contact between the subunits. The arrangement of the helices of the globular part resembles the globin fold: 59 equivalent C alpha-atoms have a root-mean-square deviation of 2 X 9 A. The chromophores attached to cystein 84 of the alpha- and beta-subunits are topologically equivalent to the haem. All three chromophores of C-phycocyanin, open-chain tetrapyrroles, are in an extended conformation. alpha 84 and beta 84 are attached to helix E (globin nomenclature), beta 155 is linked to the G--H loop. The shortest centre-to-centre distance between chromophores in trimer is 22 A.

Molecular structure of the bilin binding protein (BBP) from Pieris brassicae after refinement at 2.0 Å resolution

The bilin binding protein (BBP) from the insect Pieris brassicae has been analysed for amino acid sequence, spectral properties and three-dimensional structure. The crystal structure that had been determined by isomorphous replacement has been refined at 2.0 A (1 A = 0.1 nm) resolution to an R-value of 0.20. The asymmetric unit contains four independent subunits of BBP. The co-ordinate differences are 0.25 A, in accord with the estimated error in co-ordinates. The polypeptide chain fold is characterized by an eight-stranded barrel. The connecting loops splay out at the upper end of the barrel and open it, whilst the lower end is closed. The overall shape resembles a calyx. The biliverdin IX gamma chromophore is located in a central cleft at the upper end of the barrel. The bilatriene moiety is in cyclic helical geometry with configuration Z,Z,Z and conformation syn,syn,syn. The geometry is in accord with the spectral properties and permits a correlation between sign of the circular dichroism bands and sense of the bilatriene helices. The fold of BBP is related to retinol binding protein (RBP), as had been recognized in the preliminary analysis, although the amino acid sequences of RBP and BBP show only 10% homology. There are large differences in the loops at the upper end of the barrel, whilst the segments of the centre and the lower end of the barrel superimpose closely. The ligands of BBP and RBP, biliverdin and retinol, respectively, are also similarly located.

STRUCTURE AND FUNCTION OF LIGHT-HARVESTING COMPLEXES AND THEIR POLYPEPTIDES

The light reactions of photosynthesis are characterized by the regulated joint action of the antenna complexes (specialized in photon absorption and in energy transfer by mobile excitons) and the reaction centers (structured for exciton trapping and charge separation). In accordance with these primary reactions a basic characteristic of the photosynthetic apparatus of bacteria, cyanobacteria, algae and higher plants is the large light-harvesting antenna with its multitude of pigment molecules (BChl, bilines, Chl, carotenoids)* and the special pair of chlorophyll molecules or the other functional ligands of the reaction center. This cooperative system with a very low degree of energy dissipation and a high energy absorption (> 90%) requires a highly ordered antenna-RC system. Numerous biophysical studies and theoretical considerations have treated the function of the antenna complexes and the reaction center. In the course of these it was generally admitted that complete understanding of the function of these pigment-protein complexes can only be achieved with detailed knowledge of their structure and molecular organization. The tendency to concentrate increasingly on the biochemical characterization and structure analysis of these complexes, which already appeared in earlier reviews of these series (Cogdell and Valentine, 1983; MacColl, 1982; Barber, 1979; Thornber et al. , 1979), has become yet more marked in recent years. The detailed biochemical analysis also allowed a better identification of the various pigment protein complexes, which is essential for their isolation. From the biochemical and structural characterization of the pigment protein complexes of light harvesting antennae, the following general features were obtained: (1) The pigment molecules are specifically bound

Refined three-dimensional structure of phycoerythrocyanin from the cyanobacterium Mastigocladus laminosus at 2.7 Å

The structure of the phycobiliprotein phycoerythrocyanin from the thermophilic cyanobacterium Mastigocladus laminosus has been determined at 2.7 A resolution by X-ray diffraction methods on the basis of the molecular model of C-phycocyanin from the same organism. Hexagonal phycoerythrocyanin crystals of space group P6(3) with cell constants a = b = 156.86 A, c = 40.39 A, alpha = beta = 90 degrees, gamma = 120 degrees are almost isomorphous to C-phycocyanin crystals. The crystal structure has been refined by energy-restrained crystallographic refinement and model building. The conventional crystallographic R-factor of the final model was 19.2% with data to 2.7 A resolution. In phycoerythrocyanin, the three (alpha beta)-subunits are arranged around a 3-fold symmetry axis, as in C-phycocyanin. The two structures are very similar. After superposition, the 162 C alpha atoms of the alpha-subunit have a mean difference of 0.71 A and the 171 C alpha atoms of the beta-subunit differ by 0.51 A. The stereochemistry of the chiral atoms in the phycobiliviolin chromophore A84 is C(31)-R, C(4)-S. The configuration of the chromophore is C(10)-Z, C(15)-Z and the conformation C(5)-anti, C(9)-syn and C(14)-anti like the phycocyanobilin chromophores in phycoerythrocyanin and C-phycocyanin.

Structure and Organization of Purple Bacterial Antenna Complexes

This chapter presents a comprehensive overview of what is currently known about the structure of purple bacterial antenna complexes, with special emphasis upon modeling their three-dimensional structure and intramembrane organization.

Functional assignment of chromophores and energy transfer in C phycocyanin isolated from the thermophilic cyanobacterium Mastigocladus laminosus

The optical characteristics and pathway of energy transfer in the C phycocyanin trimer isolated from the thermophilic cyanobacterium Mastigocladus laminosus were investigated at steady state by absorption, circular dichroism, fluorescence and fluorescence polarization spectroscopy. Based on the comparison of optical data with the 3-dimensional structure of the C-phycocyanin trimer determined by X-ray analysis (Schirmer, T., Bode, W., Huber, R., Sidler, W. and Zuber, H. (1984) in Proceedings of the Symposium on Optical Properties and Structure of Tetrapyrroles, (Blauer, G. and Sund, M., eds.), pp. 445–449, Walter de Gruyter, Berlin, and (1985) J. Mol. Biol. 184, 257–277), the functional assignment of three types of chromophore was established. An α subunit has an s chromophore and the chromophores at the positions 84 and 155 in the amino acid sequence of the β subunit are assigned as f and s chromophores, respectively. In the C phycocyanin trimer energy transfer occurs from the α chromophore in one monomer to the βf chromophore in an adjacent monomer, and from the βs chromophore to the βf chromophore in the same monomer. The direction of energy flow is from the outside to the inside of the trimer, where the locus for the binding of a colourless polypeptide is postulated. In the phycobilisomes the energy concentrated at the βf chromophores might be transferred toward the allophycocyanin core mainly by the βf chromophores in the phycocyanin rods.

The complete amino acid sequence of the single light harvesting protein from chromatophores of Rhodospirillium rubrum G-9+

Pigment-free preparations of the single light-harvesting protein from the purple photosynthetic bacterium Rhodospirillum rubrum have been reported both from the antenna bacteriochlorophyll-protein complex and directly from chromatophores by chloroform/methanol extraction [ 1,2]. Molecular mass determinations yielded values of 9000-l 9 000 [ l-4 1. Preliminary sequence information was obtained from a few very small fragments of limited acid-treated LHP from wildtype R. rubrum [5]. N-Terminal amino acid sequence data have been published for LHP from the carotenoidless mutant R. r&urn G-9+, which was found to be blocked, presumably by a formyl group [6]. Aside from the elucidation of the initial N-termin’al sequences of the 10 000 Mr and the 8000 J4, polypeptides from the light-harvesting B800-850 complex from Rhodopseudomonas capsulata strain Y5 [7], this was the first report on extensive N-terminal sequence data of a LHP from a purple photosynthetic bacterium. Here we present the complete amino acid sequence of LHP from R. rubrum G-9+, thus opening the way for progress in localizing it within the chromatophore membrane and for studying its in vivo aggregational state in the BChl-protein complex. On the basis of the amino acid sequence, LHP consists of 52 amino acid residues yielding 6106 Mr. Of particular interest

The hydrophobic anchor of small-intestinal sucrase—isomaltase: N-Terminal sequence of the isomaltase subunit

The sucrase-isomaltase complex (SI), a glycoprotein of two subunits of app. mol. wt 140 OOO160 000 each, is an intrinsic protein of the smallintestinal brush border membrane [1,2]. The catalytic centres are fully accessible from the lumen, and the bulk of the protein mass probably protudes from the outer, luminal surface of the membrane ([3,4], reviewed [5,6]). Several ectoproteins have a linear domain structure near the C-terminal which encompasses the cytoplasmic and the intramembraneous regions (e.g. [7]; reviewed [S]). Small intestinal SI, instead, is anchored to the brush border membrane via a segment at the N-terminal region of one of the subunits (i.e., isomaltase) [2]. A significant interaction of the C-terminal regions with the membrane fabric could be ruled out. This different mode of anchoring of an intrinsic protein to a plasma membrane called for additional events to be postulated in the biosynthesis and/or the insertion process(es) of SI. We report here a partial amino acid sequence of the N-terminal region of the isomaltase subunit. It includes an extremely hydrophobic sequence, which agrees with and supports the conclusion reached

Isolation and characterization of cDNA clones encoding the 17.9 and 8.1 kDa subunits of Photosystem I from Chlamydomonas reinhardtii

AbstractcDNA clones encoding two Photosystem I subunits of Chlamydomonas reinhardtii with apparent molecular masses of 18 and 11 kDa (thylakoid polypeptides 21 and 30; P21 and P30 respectively) were isolated using oligonucleotides, the sequences of which were deduced from the N-terminal amino acid sequences of the proteins. The cDNAs were sequenced and used to probe Southern and Northern blots. The Southern blot analysis indicates that both proteins are encoded by single-copy genes. The mRNA sizes of the two components are 1400 and 740 nucleotides, respectively. Comparison between the open reading frames of the cDNAs and the N-terminal amino acid sequences of the proteins indicates that the molecular masses of the mature proteins are 17.9 (P21) and 8.1 kDa (P30). Analysis of the deduced protein sequences predicts that both subunits are extrinsic membrane proteins with net positive charges. The amino acid sequences of the transit peptides suggest that P21 and P30 are routed towards the lumenal and stromal sides of the thylakoid membranes, respectively.

Structure, function and organization of antenna polypeptides and antenna complexes from the three families of Rhodospirillaneae.

Comparative primary structural analysis of polypeptides from antenna complexes from species of the three families of Rhodospirillaneae indicates the structural principles responsible for the formation of spectrally distinct light-harvesting complexes. In many of the characterized antenna systems the basic structural minimal unit is an alpha/beta polypeptide pair. Specific clusters of amino acid residues, in particular aromatic residues in the C-terminal domain, identify the antenna polypeptides to specific types of antenna systems, such as B880 (strong circular dichroism (CD)), B870 (weak CD), B800-850 (high), B800-850 (low) or B800-820. The core complex B880 (B1020) of species from Ectothiorhodospiraceae and Chromatiaceae apparently consists of four (alpha 1 alpha 2 beta 1 beta 2) or three (2 alpha beta 1 beta 2) chemically dissimilar antenna polypeptides respectively. There is good evidence that the so-called variable antenna complexes, such as the B800-850 (high), B800-850 (low) or B800-820 of Rp. acidophila, Rp. palustris and Cr. vinosum, are comprised of multiple forms of peripheral light-harvesting polypeptides. Structural similarities between prokaryotic and eukaryotic antenna polypeptides are discussed in terms of similar pigment organization. The structural basis for the strict organization of pigment molecules (bacteriochlorophyll (BChl) cluster) in the antenna system of purple bacteria is the hierarchical organization of the alpha- and beta-antenna polypeptides within and between the antenna complexes. On the basis of the three-domain structure of the antenna polypeptides with the central hydrophobic domain, forming a transmembrane alpha helix, possible arrangements of the antenna polypeptides in the three-dimensional structure of core and peripheral antenna complexes are discussed. Important structural and functional features of these polypeptides and therefore of the BChl cluster are the alpha/beta heterodimers, the alpha 2 beta 2 basic units and cyclic arrangements of these basic units. Equally important for the formation of the antenna complexes or the entire antenna are polypeptide-polypeptide, pigment-pigment and pigment-polypeptide interactions.