Jürgen Marquardt

Isolation and characterization of biliprotein aggregates from Acaryochloris marina, a Prochloron-like prokaryote containing mainly chlorophyll d

Phycobiliprotein aggregates were isolated from the prokaryote Acaryochloris marina, containing chlorophyll d as major pigment. In the electron microscope the biliprotein aggregates appear as rod‐shaped structures of 26.0×11.3 nm, composed of four ring‐shaped subunits 5.8 nm thick and 11.7 nm in diameter. Spectral data indicate that the aggregates contain two types of biliproteins: phycocyanin and an allophycocyanin‐type pigment, with very efficient energy transfer from the phycocyanin‐ to allophycocyanin‐type constituent. The chromophore‐binding polypeptides of the pigments have apparent molecular masses of 16.2 and 17.4 kDa. They crossreact with antibodies against phycocyanin and allophycocyanin from a red alga.

Molecular structure, localization and function of biliproteins in the chlorophyll a/d containing oxygenic photosynthetic prokaryote Acaryochloris marina

We investigated the localization, structure and function of the biliproteins of the oxygenic photosynthetic prokaryote Acaryochloris marina, the sole organism known to date that contains chlorophyll d as the predominant photosynthetic pigment. The biliproteins were isolated by means of sucrose gradient centrifugation, ion exchange and gel filtration chromatography. Up to six biliprotein subunits in a molecular mass range of 15.5-18.4 kDa were found that cross-reacted with antibodies raised against phycocyanin or allophycocyanin from a red alga. N-Terminal sequences of the alpha- and beta-subunits of phycocyanin showed high homogeneity to those of cyanobacteria and red algae, but not to those of cryptomonads. As shown by electron microscopy, the native biliprotein aggregates are organized as rod-shaped structures and located on the cytoplasmic side of the thylakoid membranes predominantly in unstacked thylakoid regions. Biochemical and spectroscopic analysis revealed that they consist of four hexameric units, some of which are composed of phycocyanin alone, others of phycocyanin together with allophycocyanin. Spectroscopic analysis of isolated photosynthetic reaction center complexes demonstrated that the biliproteins are physically attached to the photosystem II complexes, transferring light energy to the photosystem II reaction center chlorophyll d with high efficiency.

The membrane-intrinsic light-harvesting complex of the red alga Galdieria sulphuraria (formerly Cyanidium caldarium): biochemical and immunochemical characterization

Abstract The membrane-intrinsic light-harvesting complex of the red alga Galdieria sulphuraria (formerly Cyanidium caldarium ) could be isolated by gel-electrophoresis as a green band with an apparent molecular mass of about 20 kDa. The band had a long-wavelength absorption maximum at 672 nm and a fluorescence maximum (77 K) at 680 nm and reacted with an antibody against light-harvesting proteins of higher plant Photosystem I. Screening of thylakoid membranes with antisera directed against various chlorophyll a/b and chlorophyll a/c light-harvesting proteins indicated the existence of at least 4 distinct light-harvesting polypeptides with apparent molecular masses between 17 and 20 kDa. Isolation of Photosystem I and of a fraction enriched in Photosystem II showed that these polypeptides are exclusively bound to Photosystem I, thus forming a holocomplex which binds at least 205 molecules of chlorophyll a , and 33 and 37 molecules of zeaxanthin and β-carotene, respectively. Additionally, there is some evidence for the existence of a second Photosystem I pool without light-harvesting complexes. In-vitro translation experiments showed that at least two of the five polypeptides which constitute the membrane-intrinsic light-harvesting complex of Galdieria sulphuraria are translated from the poly(A)-enriched RNA fraction. They could be immunoprecipitated as preproteins being 3 to 4 kDa larger in size than the mature polypeptides.

Effects of carotenoid-depletion on the photosynthetic apparatus of a Galdieria sulphuraria (rhodophyta) strain that retains its photosynthetic apparatus in the dark

Summary A Galdieria sulphuraria (Galdieri) Merola strain that retains its photosynthetic apparatus in the dark was treated with the inhibitor of carotenoid biosynthesis, norflurazon, under autotrophic and heterotrophic conditions. Under autotrophic conditions cultures were not able to grow. The chlorophyll content of the cells decreased constantly and a partial decomposition of their chloroplast structure could be observed. In the dark, under heterotrophic conditions, the inhibitor caused a nearly total loss of carotenoids, a reduction of the chlorophyll content per cell, a slight increase of the ratio of phycocyanin to chlorophyll and an accumulation of a pheophorbide a -like pigment. The ultrastructure of the algae, however, was not affected. The polypeptides of the photosystem II core-complex and of the light-harvesting complex of photosystem I were dramatically reduced, indicating that carotenoids are essential for their assembly. The core-complex of photosystem I was present. Its fluorescence maximum, however, was blue-shifted from 725 to 717nm. When excited at 580 nm norflurazon-treated cells exhibited a fluorescence maximum at 686 nm, indicating that phycobilisomes are energetically neither connected with photosystem II nor with photosystem I. Nevertheless, phycobilisomes of norflurazon-treated and control algae were not distinguishable.

Intron–exon structure and gene copy number of a gene encoding for a membrane-intrinsic light-harvesting polypeptide of the red alga Galdieria sulphuraria

Genes for light-harvesting proteins (lhc genes) of higher plants are well examined. However, little is known about the corresponding genes of algae, although this knowledge might give valuable information about the evolution of photosynthetic antennae. In the case of rhodophytes only two cDNA sequences from a single organism, Porphyridium cruentum, have been published. Here we describe an additional sequence from another species, the thermo-acidophilic red alga Galdieria sulphuraria. For the first time also a genomic sequence for a red algal lhc gene is presented. From a cDNA library of G. sulphuraria we isolated a clone containing an open reading frame for a protein of 302 amino acids with a deduced molecular mass of 33.86kDa. It shares major structural features with eukaryotic light-harvesting polypeptides. A proposed cleavage site between transit peptide and mature protein gives rise to a transit peptide of 119 amino acids and a mature protein of 183 residues. Hydropathy analysis suggests that the mature protein consists of three transmembrane helices. Several amino acid residues supposed to bind chlorophyll a and chlorophyll b in higher plants are conserved. The protein shows up to 69% identity and 81% similarity to the Porphyridium polypeptides in the transmembrane helices 1 and 3. Using oligonucleotides annealing in the regions of the start and stop codons of the gene as primers, a DNA sequence was amplified from nuclear G. sulphuraria DNA by PCR. Compared with the cDNA clone, this sequence contains five additional intervening DNA strings of 50-74bp length. Four of them show typical features of spliceosomal introns with GT-AG borders, and the fifth differs by starting with GC. Three of the supposed introns are located in similar positions as introns of higher plant light-harvesting proteins. Southern blotting and hybridization experiments indicate that G. sulphuraria contains at least three copies of this gene.

The gene family coding for the light-harvesting polypeptides of Photosystem I of the red alga Galdieria sulphuraria*

Recently [Marquardt et al. (2000) Gene 255: 257–265], we isolated a gene encoding a polypeptide of the light-harvesting complex of Photosystem I (LHC I) of the red alga Galdieria sulphuraria. By screening a G. sulphuraria cDNA library with a DNA probe coding for the conserved first transmembrane helix of this protein we isolated four additional genes coding for LHC I polypeptides. The deduced preproteins had calculated molecular masses of 24.6–25.6 kDa and isoelectric points of 8.09–9.82. N-terminal sequencing of a LHC I polypeptide isolated by gel electrophoresis allowed us to determine the cleavage site of the transit peptide of one of the deduced polypeptides. The mature protein has a calculated molecular mass of 20.6 kDa and an isoelectric point of 7.76. The genes were amplified from nuclear G. sulphuraria DNA by polymerase chain reaction (PCR) using oligonucleotides annealing in the regions of the start and stop codons as primers. All genomic sequences were 80–300 base pairs longer than the PCR products obtained from the respective cDNA clones, pointing to the existence of 1–5 introns per gene. The G. sulphuraria genes form a homogeneous gene family with overall pairwise amino acid identities of 46.0–56.6%. Homology to two diatom, one cryptophytic and two higher plant light-harvesting polypeptides was lower with pairwise identities of 21.1–34.1%. Only one diatom polypeptide showed a higher degree of identity of up to −39.3%.

Porphyridium purpureum (Rhodophyta) from red and green light: characterization of photosystem I and determination of in situ fluorescence spectra of the photosystems

Abstract Recent investigations have shown the existence of a light-harvesting complex (LHC) of photosystem I (PSI) with a fluorescence maximum at 680 nm in the red alga Porphyridium purpureum (Wolfe et al., Nature, 367 (1994) 566–568). We have attempted to determine whether the composition of this red algal LHC is invariable or subject to light acclimatization and whether the 680–690 nm fluorescence band of intact algae, which is usually ascribed to photosystem II (PSII), could be partially due to the LHC of PSI. For this purpose, we characterized PSI core complexes and PSI holocomplexes consisting of core and LHC from algae grown under red or green light conditions and calculated the in situ spectra of the photosystems by normalizing the thylakoid spectra of red and green light algae to an identical PSI or PSII concentration. The composition of the LHC seems to be independent of the light quality, since no significant difference was found in the antenna size and the pigment composition of the PSI holocomplexes from red and green light algae; both holocomplex preparations contained 150–160 chlorophylls, 21–22 zeaxanthins and 23–25 β-carotenes per reaction centre, while the core complexes contained approximately 90 chlorophylls, 15 carotenes and no zeaxanthin. Both holocomplex preparations had a fluoroscence maximum (77 K) at 718 nm. A short-wavelength emission band, as reported by Wolfe et al., was not found in the spectra of the isolated PSI complexes or in the PSI spectrum calculated from thylakoid spectra. This indicates that the LHC does not contribute to the fluorescence spectrum of intact cells, but transfers energy very effectively to the PSI core complex.

The epizoic diatom community on four bryozoan species from Helgoland (German Bight, North Sea)

Abstract. The composition of the diatom community on the bryozoans Electra pilosa, Membranipora membranacea, Flustra foliacea, and Alcyonidium gelatinosum from the German Bight was studied by light and scanning electron microscopy. In total, members of 26 diatom genera were found, with Cocconeis, Tabularia, Licmophora, Amphora, and Navicula being the most abundant. The amount and the composition of the diatom covering seem to be typical for single bryozoan species. Electra pilosa and Membranipora membranacea showed a rather dense covering with 71–547 cells/mm2 and 77–110 cells/mm2, respectively. The most prominent genus on Electra pilosa was Cocconeis, reaching up to 58% of all diatoms in one sample, followed by Navicula, Tabularia and Amphora. The most abundant genera on Membranipora membranacea were Tabularia and Licmophora, making up almost 70% of all diatoms in one sample, followed by Navicula, Cocconeis and Amphora. The diatom composition was very stable on all Electra samples, but varied on Membranipora samples. With <1–27 cells/mm2, diatoms were much less abundant on Alcyonidium gelatinosum. Members of the genera Tabularia and Navicula were the most frequently found benthic diatoms, whereas the planktonic forms Coscinodiscus, Cyclotella, and Thalassiosira made up 35% of the diatoms. On Flustra foliacea, diatoms were virtually absent, with fewer than 5 cells/mm2. The low diatom numbers are probably due to toxic metabolites produced by the host. The same may be true for Alcyonidium gelatinosum, but here they might also be a consequence of the surface properties of the bryozoan.

The taxonomic position of Chlamydomyxa labyrinthuloides

Chlamydomyxa labyrinthuloides is a heterokont alga known since the last century. It lives on Sphagnum and other water plants as aplanospores or plasmodia. We have investigated the taxonomic position of Chlamydomyxa labyrinthuloides by combining results from morphological studies, pigment analyses and a molecular phylogenetic analysis of the small subunit (SSU) rRNA gene. Chlamydomyxa labyrinthuloides shares morphological features with xanthophytes and chrysophytes, whereas pigment composition indicates a grouping with the phaeophytes, raphidophytes and chrysophytes. The sequence of the SSU rRNA gene and its phylogenetic reconstruction unambiguously demonstrate that Chlamydomyxa labyrinthuloides is related to the chrysophytes.

THE BIG TRAIL: MANY MIGRATE AT THE EXPENSE OF A FEW

Wadden Sea inhabiting diatoms often show a patchy distribution on sediment surfaces with areas of higher and lower cell densities. The factors causing patchiness have not been clearly worked out and are under debate. Even the mode of reaching the sediment surface is almost completely unknown. The observations indicate that trails of individual diatoms might be used by others for upward directed migration and offer another plausible explanation for the existence of surfaces areas of higher and lower cell densities in general.