Biotransformation of 1-naphthalenesulfonic acid by the green alga Scenedesmus obliquus

Under sulfate limitation, axenic batch cultures of the green alga Scenedesmus obliquus metabolized 1-naphthalenesulfonic acid and partially used the sulfonate as a source of sulfur. The main metabolite, 1-hydroxy-2-naphthalenesulfonic acid, which was not metabolized further in the algal culture, was formed by hydroxylation of the substrate in position 1 and by migration of the sulfonic acid group to position 2 of the naphthalene ring (NIH shift). A smaller amount of 1-naphthalenesulfonic acid was desulfonated. The resulting 1-naphthol was mostly transformed into 1-naphthyl β-d-glucopyranoside. Received: 27 March 1996 / Revision received: 18 October 1996 / Accepted: 30 October 1996     

Effects of CO2 concentrations on the freshwater microalgae, Chlamydomonas reinhardtii, Chlorella pyrenoidosa and Scenedesmus obliquus (Chlorophyta)

In order to investigate the possible impacts of increased atmospheric CO₂ levels on algal growth and photosynthesis, the influence of CO₂ concentration was tested on three planktonic algae (Chlamydomonas reinhardtii, Chlorella pyrenoidosa, and Scenedesmus obliquus). Increased CO₂ concentration enhanced significantly the growth rate of all three species. Specific growth rates reached maximal values at 30, 100, and 60 M CO₂ in C. reinhardtii, C. pyrenoidosa, and S. obliquus, respectively. Such significant enhancement of growth rate with enriched CO₂ was also confirmed at different levels of inorganic N and P, being more profound at limiting levels of N inC. pyrenoidosa and P in S. obliquus. The maximal rates of net photosynthesis, photosynthetic efficiency and light-saturating point increased significantly (p < 0.05) in high-CO₂-grown cells. Elevation of the CO₂ levels in cultures enhanced the photoinhibition of C. reinhardtii, but reduced that of C. pyrenoidosa and S. obliquus when exposed to high photon flux density. The photoinhibited cells recovered to some extent (from 71% to 99%) when placed under dim light or in darkness, with better recovery in high-CO₂-grownC. pyrenoidosa and S. obliquus. Although pH and pCO₂ effects cannot be distinguished from this study, it can be concluded that increased CO₂ concentrations with decreased pH could affect the growth rate and photosynthetic physiology of C. reinhardtii, C. pyrenoidosa, and S. obliquus.     

Alkylsulfonic acids and some S-containing detergents as sulfur sources for growth of Chlorella fusca

Chlorella fusca can utilize the following substances as sole sulfur sources for growth: C₁ to C₈ n-alkane-1-sulfonates, linear alkylbenzenes sulfonates (LAS), -sulfonated fatty acid esters, polyethylene glycol sulfate and alkylsulfates. Good sulfur sources are alkylsulfonic acids, which are comparable to sulfate. Ethanesulfonic acid was used for comparison of the growth on sulfate and on a sulfonic acid, because best growth was achieved on this C₂-sulfonic acid.Growth data of Chlorella on the enviromental important detergents linear alkylbenzene sulfonic acids, -sulfonated fatty acid methylester, Texapon and Sulfopon are presented. So far only microorganisms have been discussed as a source for degradation of sulfonic acids and detergents. It is suggested that green algae could be of similar importance for the biodegradation of these compounds.Abbreviations LAS Linear alkylbenzene sulfonate - ES -sulfonated fatty acid methylester - DTE dithiocrythritol     

Microalgae as a raw material for biofuels production

Biofuels demand is unquestionable in order to reduce gaseous emissions (fossil CO₂, nitrogen and sulfur oxides) and their purported greenhouse, climatic changes and global warming effects, to face the frequent oil supply crises, as a way to help non-fossil fuel producer countries to reduce energy dependence, contributing to security of supply, promoting environmental sustainability and meeting the EU target of at least of 10% biofuels in the transport sector by 2020. Biodiesel is usually produced from oleaginous crops, such as rapeseed, soybean, sunflower and palm. However, the use of microalgae can be a suitable alternative feedstock for next generation biofuels because certain species contain high amounts of oil, which could be extracted, processed and refined into transportation fuels, using currently available technology; they have fast growth rate, permit the use of non-arable land and non-potable water, use far less water and do not displace food crops cultures; their production is not seasonal and they can be harvested daily. The screening of microalgae (Chlorella vulgaris, Spirulina maxima, Nannochloropsis sp., Neochloris oleabundans, Scenedesmus obliquus and Dunaliella tertiolecta) was done in order to choose the best one(s), in terms of quantity and quality as oil source for biofuel production. Neochloris oleabundans (fresh water microalga) and Nannochloropsis sp. (marine microalga) proved to be suitable as raw materials for biofuel production, due to their high oil content (29.0 and 28.7%, respectively). Both microalgae, when grown under nitrogen shortage, show a great increase (~50%) in oil quantity. If the purpose is to produce biodiesel only from one species, Scenedesmus obliquus presents the most adequate fatty acid profile, namely in terms of linolenic and other polyunsaturated fatty acids. However, the microalgae Neochloris oleabundans, Nannochloropsis sp. and Dunaliella tertiolecta can also be used if associated with other microalgal oils and/or vegetable oils.     

Carbon dioxide fixation by Chlorella kessleri, C. vulgaris, Scenedesmus obliquus and Spirulina sp. cultivated in flasks and vertical tubular photobioreactors

CO₂ at different concentrations were added to cultures of the eukaryotic microalgae, Chlorella kessleri, C. vulgaris and Scenedesmus obliquus, and the prokaryotic cyanobacterium, Spirulina sp., growing in flasks and in a photobioreactor. In each case, the best kinetics and carbon fixation rate were with a vertical tubular photobioreactor. Overall, Spirulina sp. had the highest rates. Spirulina sp., Sc. obliquus and C. vulgaris could grow with up to 18% CO₂.     

Effects of the novel allelochemical ethyl 2-methylacetoacetate from the reed (<Emphasis Type="Italic">Phragmitis australis</Emphasis> Trin) on the growth of several common species of green algae

Bioassays were performed to investigate the effects of the novel allelochemical, ethyl 2-methylacetoacetate (EMA), isolated from the reed (Phragmitis australis) on the growth of three common species of algae; Scenedesmus obliquus, Selenastrum capricornutum and Chlamydomonas reinhardtii. The results demonstrated that EMA has three quite different types of effect on these three species of algae. The growth of S. capricornutum was significantly inhibited by EMA during the whole cultivation period. The EC₅₀ values of EMA on S. capricornutum was 0.6 mg L⁻¹(7 days). However, the inhibitory effect of EMA on S. obliquus was apparent during the first 4 days of batch cultivation and then the inhibitory effect disappeared, and a stimulating effect was observed instead. The EC₅₀ value of EMA on S. obliquus was 0.43 mg L⁻¹(4 days). In addition, following the addition of EMA, the cells of S. obliquus and S. capricornutum became significantly larger than the normal untreated one and the algal cells changed morphologically. The microstructure of the algal cells was disrupted by the addition of EMA. There was no significant inhibition of the growth of C. reinhardtii by EMA, but cell motility was affected.     

A culture method for microalgal forms using two-tier vessel providing carbon-dioxide environment: studies on growth and carotenoid production

A culture method was developed for photoautotrophic culture of Haematococcus pluvialis, Chlorella vulgaris, Scenedesmus obliquus, Spirulina platensis, Nostoc and Stigonema in a two-tier flask consisting of nutrient media in the upper chamber and CO₂ generating buffer mixture (KHCO₃/K₂CO₃) in the lower chamber. The concentration of buffer mixture was varied to obtain desired levels of CO₂. CO₂ at 2.0% (v/v) level enhanced growth and chlorophyll content over control cultures (without CO₂ supplementation) in all microalgal species. Haematococcus pluvialis culture in BBM and KM1 media showed 6.71- and 2.07-fold increase in biomass yields with astaxanthin productivity at 7.26 and 7.48 mg l^–1 level respectively. CO₂ supplementation to C. vulgaris and S. obliquus cultures resulted in 5.97- and 7.30-folds increase in biomass with 2–3 fold increase in chlorophyll and carotenoid contents over their respective controls. Similarly 2–3 fold increase in chlorophyll and carotenoid contents were observed in Sp. platensis, Nostoc and Stigonema spp. This culture methodology will provide information on CO₂ requirement for growth of algae and metabolite production and also facilitates studies on the influence of light and temperature conditions.     

Allelopathic effects of the submerged macrophyte <Emphasis Type="Italic">Potamogeton malaianus</Emphasis> on <Emphasis Type="Italic">Scenedesmus obliquus</Emphasis>

Allelopathic effects of the submerged macrophyte Potamogeton malaianus on Scenedesmus obliquus were assessed using a two-phase approach under controlled laboratory conditions. In the co-culture experiment (phase І), the growth of S. obliquus at two different initial cell densities was significantly inhibited by P. malaianus. Moreover, the growth inhibition was dependent on the biomass density of P. malaianus. Antioxidant enzymes (SOD, CAT and POD), MDA, APA, total soluble protein, protein electrophoretic pattern and morphology of S. obliquus were determined after the co-culture experiment was terminated. The activities of SOD, CAT, POD and APA at the low initial cell density were stimulated, the contents of MDA and total soluble protein were increased, and some special protein bands disappeared in P. malaianus treatments. The macrophyte had no effect on the activities of SOD and APA at the high initial cell density, but significantly influenced other physiological parameters of S. obliquus with the increase of biomass density. The morphology of S. obliquus showed no difference in the macrophyte treatments and the controls, and the cultures were dominated by 4-celled coenobia. The results indicated P. malaianus had significant allelopathic effects on the growth and physiological processes of S. obliquus. Moreover, the allelopathic effects depended on initial algal cell density, biomass density of the macrophyte, and their interaction. In the experiment of P. malaianus culture filtrates (phase II), filtrates from combined culture of plant and S. obliquus at the low initial cell density exhibited no apparent growth inhibitory effect on S. obliquus. The result showed that initial addition of growth-inhibiting plant filtrates had no allelopathic effect on S. obliquus. We concluded that the allelopathic effects on S. obliquus were found in the presence of P. malaianus, but not in P. malaianus filtrates. However, the absence of allelopathic effect on S. obliquus might be due to the very low concentrations of allelochemicals in the filtrates. Handling editor: S. M. Thomas     

Relationship between biosynthesis of carotenoids and increasing complexity of photosystem I in mutant C-6D of Scenedesmus obtiquus

Dark-grown cells of the pigment mutant C-6D of Scenedesmus obliquus, strain D₃ (Gaffron 1939), contain only chlorophyll (Chl) a and carotenoid precursors. In these cells a functioning photosystem I (PSI) of basic structure was characterised by a high PSI activity and a low Chl/P700 ratio. The reaction-center complex of PSI (CPI) was shown to exist in the dark-grown cells. These findings demonstrate that the assembly of the core complex of PSI and its function are independent of the presence of carotenoids. Upon illumination, carotenoids, Ch1 b and additional Chl a were synthesized. Newly formed -carotene was shown by pigment analysis using high-performance liquid chromatography (HPLC) to be incorporated into CPI. Parallel to this process a shift of the long-wavelength fluorescence emission of PSI from 712–714 to 718–719 nm was observed. In the later stages of chloroplast differentiation, when xanthophylls and Chl b were synthesized, a higher-molecular-weight complex of PSI (CPIa) could be isolated. Pigment analysis demonstrated that CPIa contained xanthophylls and Chl b in addition to Chl a and -carotene. This indicates the formation of a light-harvesting antenna closely associated with PSI (LHCI). The addition of an LHCI to the reaction-center complex of PSI caused an increase in the absorption cross-section of PSI as shown by action spectroscopy and in-vivo fluorescence measurements. A model demonstrating the changes in the molecular organization of PSI during light-induced carotenoid biosynthesis in mutant C-6D of Scenedesmus obliquus is presented.Abbreviations Chl chlorophyll - CP chlorophyll-protein complex - LHC light-harvesting complex - HPLC high-performance liquid chromatography - PSI, II photosystem I, II - PAGE polyacrylamide gel electrophoresis This work was supported by the Deutsche Forschungsgemeinschaft and a scholarship of the Studienstiftung des deutschen Volkes to S. Römer. We thank Ms. K. Bölte for technical assistance and Mr. H. Becker for drafting the figures.     

Luminescence decay kinetics in relation to quenching and stimulation of dark fluorescence from high and low CO2 adapted cells of Scenedesmus obliquus and Chlamydomonas reinhardtii

Two green algal species, Chlamydomonas reinhardtii and Scenedesmus obliquus, exhibited a relative maximum during the decay of luminescence, when adapted to low CO₂ conditions that was not observed in high CO₂ adapted cells.From the kinetics of transient changes in the level of dark fluorescence, after illumination and parallel to the luminescence maxima, it was concluded that the maximum in Scenedesmus was mainly related to a decrease in nonphotochemical quenching, whereas in Chlamydomonas the maximum was mainly related to a dark reduction of the primary PS II acceptor Q_A.ATP/ADP ratios from low CO₂ adapted Scenedesmus showed transient high levels after a dark/light transition that was not observed in high CO₂ adapted cells. After 30 s of illumination the ATP/ADP ratios however stabilized at the same steady state level as in high CO₂ adapted cells.Dark addition of HCO₃ ^- to low CO₂ adapted cells of Chlamydomonas resulted in a rapid transient quenching of luminescence that was not observed in low CO₂ adapted cells of neither species.It is concluded that the luminescence maxima present in both low CO₂ adapted Scenedesmus and Chlamydomonas reflect adaptation of the cells to low CO₂ conditions. It is further suggested that the difference in mechanistic origin of luminescence maxima in the two species reflects differences in adaptation.Abbreviations ADP adenosine-diphosphate - ATP adenosine-triphosphate - C_i inorganic carbon - F_D dark fluorescence recorded under dark adapted conditions - F₀ fluorescence with all reaction centers open - FV variable fluorescence - PS I photosystem I - PS II photosystem II - Q_A the first quinone acceptor of PS II     

Possible toxic effects on Daphnia resulting from the green alga Scenedesmus obliquus

We cultured individuals of two Daphnia species and their hybrid on two different algae, Scenedesmus obliquus and Chlamydomonas globosa, in different concentrations. Our results suggest that culture conditions of S. obliquus can be such that the algal cells become toxic to Daphnia     

Interactions of glycolate-, HCO 3 - -, Cl--, and H+-balance of Scenedesmus obliquus

Excretion and absorption of glycolate by young cells of Scenedesmus obliquus (Turp.) Krüger strain D₃ grown synchronously with 2% CO₂ was compared after no pretreatment with air (CO₂-adapted) or after a 2 h adaptation to normal air (0.03% CO₂) (air-adapted). At 21% O₂, excretion occurred only from CO₂-adapted cells at high pH (pH 8.0). Under conditions where no excretion occurred, external glycolate (0.2 mM) was taken up by both air-and CO₂-adapted cells at a much faster rate at pH 5 than at pH 8. The uptake was accompanied by an apparent stoichiometric uptake of H⁺. CO₂-adapted algae exhibited high uptake rates that were even higher in the dark than in the light. Air-adapted algae showed high uptake rates in the light but only minimal uptake in the dark. The uptake rate was decreased to about 1/3 with 5% CO₂, except with CO₂-adapted cells in the light, in which a slight stimulation occurred. Cl^- ions inhibited glycolate uptake by air-adapted cells in the light; conversely, light-stimulated Cl^- uptake of these cells was inhibited by glycolate. A hypothesis is discussed according to which the internal pH regulates the uptake and release of Cl^-, HCO ₃ ^- , and glycolate.Abbreviations DCMU 3-(3,4 dichlorophenyl)-1, 1-dimethyl urea - FCCP carbonyl cyanide p-trifluoro-methoxyphenylhydrazone - HEPES 2-(4-(2-hydroxyethyl)-piperazinyl) ethanesulfonic acid - HPMS -hydroxypyridinemethanesulfonate - MES 2-morpholinoethanesulfonic acid - PCV packed cell volume     

C-nuclear magnetic resonance studies of the biosynthesis of 5-aminolevulinic acid destined for chlorophyll formation in dark-grown Scenedesmus obliquus

The ¹³C-nuclear magnetic resonance (NMR) spectra of chlorophyll a formed in dark-grown Scenedesmus obliquus (Turp.) Kützing in the presence of [1-¹³C]glutamate, [2-¹³C]- and [1-¹³C]glycineshowed that the ¹³C of glutamate was specifically incorporated into the eight-carbon atoms in the tetrapyrrole macrocycles derived from C-5 of 5-aminolevulinic acid (ALA), while the C-2 of glycine was only incorporated into the methyl carbon of the methoxycarbonyl group attached to the isocyclic ring of chlorophyll a. No specific enrichment of these nine carbon atoms was observed in the spectrum of chlorophyll a formed in the presence of [1-¹³C]-glycine. These labeling patterns provide evidence for the operation of the C₅-pathway and against the operation of the ALA synthase pathway for chlorophyll formation in darkness.     

Regeneration of the high-affinity manganese-binding site in the reaction center of an oxygen-evolution deficient mutant of Scenedesmus by protease action

The O₂-evolution deficient mutant (LF-1) of Scenedesmus obliquus inserts an unprocessed D1 protein into the thylakoid membrane and binds less than half the wild type (WT) level of Mn. LF-1 photosystem II (PS II) membrane fragments lack that part of the high-affinity Mn²⁺-binding site found in WT membranes which may be associated with histidine residues on the D1 protein (Seibert et al. 1989 Biochim Biophys Acta 974: 185–191). Hsu et al. (1987 Biochim Biophys Acta 890: 89–96) purport that the high-affinity site (characterized by competitive inhibition of DPC-supported DCIP photoreduction by M concentrations of Mn²⁺) in Mn-extracted PS II membranes is also the binding site for Mn functional in O₂ evolution. Proteases (papain, subtilisin, and carboxypeptidase A) can be used to regenerate the high-affinity Mn²⁺-binding site in LF-1 PS II membranes but not in thylakoids. Experiments with the histidine modifier, DEPC, suggest that the regenerated high-affinity Mn²⁺-binding sites produced by either subtilisin or carboxypeptidase A treatments were the same sites observed in WT membranes. However, none of the protease treatments produced LF-1 PS II membranes that could be photoactivated. Reassessment of the processing studies of Taylor et al. (1988 FEBS Lett 237: 229–233) lead us to believe that their procedure also does not result in substantial photoactivation of LF-1 PS II membranes. We conclude that (1) the unprocessed carboxyl end of the D1 protein in LF-1 is located on the lumenal side of the PS II membrane, (2) the unprocessed fragment physically obstructs or perturbs that part of the high-affinity Mn²⁺-binding site undetectable in LF-1, and (3) the D1 protein must be processed at the time of insertion into the membrane for normal O₂-evolution function to result.Abbreviations Chl chlorophyll - DCBQ 2,6-dichloro-1,4-benzoquinone - DCIP 2,6-dichlorophenol indophenol - DEPC diethylpryocarbonate - DPC 1,5-diphenylcarbazide - HEPES 4-(2-hydroxyethyl)-1-piperazine-ethanesulfonic acid - LDS-PAGE lithium dodecylsulfate polyacrylamide gel electrophoresis - LF-1 a low-fluorescent mutant of Scenedesmus obliquus - MES 4-morpholineethanesulfonic acid - PS II photosystem II - PMSF phenylmethylsulfonyl fluoride - RC photosystem II reaction center - Tris tris(hydroxymethyl)aminomethane - WT wild type Operated by the Midwest Research Institute for the U.S. Department of Energy under contract DE-AC-02-83CH10093.     

Rates of glycolate synthesis and metabolism during photosynthesis of Euglena and microalgae grown on low CO2

The rate of glycolate excretion in Euglena gracilis Z and some microalgae grown at the atmospheric level of CO₂ was determined using amino-oxyacetate (AOA). The extracellular O₂ concentration was kept at 240 M by bubbling the incubation medium with air. Glycolate, the main excretion product, was excreted by Euglena at 6 mol·h^-1·(mg chlorophyll (Chl))^-1. Excretion depended on the presence of AOA, and was saturated at 1 mM AOA. A substituted oxime formed from glyoxylate and AOA was also excreted. Bicarbonate added at 0.1 mM did not prevent the excretion of glycolate. The excretion of glycolate increased with higher O₂ concentrations in the medium, and was competitively inhibited by much higher concentrations of bicarbonate. Aminooxyacetate also caused excretion of glycolate from the green algae, Chlorella pyrenoidosa, Scenedesmus obliquus and Chlamydomonas reinhardtii grown on air, at the rates of 2–7 mol·h^-1·(mg Chl)^-1 in the presence of 0.2–0.6 mM dissolved inorganic carbon, but the cyanobacterium, Anacystis nidulans, grown in the same way did not excrete glycolate. The efficiency of the CO₂-concentrating mechanism to suppress glycolate formation is discussed on the basis of the magnitude of glycolate formation in these low-CO₂-grown cells.Abbreviations AOA aminooxyacetate - Chl chlorophyll - DIC dissolved inorganic carbon - HPLC high-pressure liquid chromatography - Rubisco ribulose-1,5-bisphosphate carboxylase/oxygenase This is the 16th paper in a series on the metabolism of glycolate in Euglena gracilis. The 15th paper is Yokota et al. (1985c)     

Purification and partial characterization of a glutamyl-tRNA synthetase from the unicellular green algaScenedesmus obliquus,mutant C-2A′

The synthesis of 5-aminolevulinic acid commences with the ligation of glutamate to a specific tRNA^Glu by a glutamyl-tRNA synthetase (E.C. 6.1.1.17) (Huang et al., 1984, Science 225, 1482–1484). The synthetase from the yellow pigment mutant C-2A of the unicellular green alga Scenedesmus obliquus was purified by sequential column chromatography on Sephacryl S-300, Blue Sepharose, phosphocellulose P11 and by fast protein liquid chromatography (FPLC) on Mono Q. After denaturing sodium dodecylsulfate (SDS)-gel electrophoresis the purified enzyme preparation revealed a single protein band with a molecular mass of 55 kDa, proving the apparent homogeneity of the glutamyl-tRNA synthetase. A molecular mass of 105 ± 10 kDa was determined for the native protein by chromatography on Sephadex G-150. From these data it can be concluded that the glutamyl-tRNA synthetase from S. obliquus is a homodimer. The purified protein is active within a pH range from 7.0 to 9.0 with a maximum activity at pH 8.0. Kinetics for the binding of glutamate to the tRNA, performed with highly purified enzyme preparations, showed a K _m value of 2.3 M ± 0.3 for glutamate.Abbreviations ALA 5-aminolevulinic acid - FPLC fast protein liquid chromatography - Glu glutamate - Hepes N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid - SDS sodium dodecylsulfate - Tricine N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]-glycine This work was supported by a grant of the Deutsche Forschungsgemeinschaft. U.C. Vothknecht is grateful for a Nachwuchs-förderungsstipendium des Landes Hessen. The authors want to thank Ms. B. Böhm, J. Gade and K. Eckhardt for skillful technical assistance. The authors also want to thank Dr. C.G. Kannangara (Carlsberg Institute, Kopenhagen, Denmark) for the donation of tRNA from barley and Dr. D. Jahn (FB Biology/Microbiology, Philipps-University, Marburg, FRG) for the tRNA^Glufrom E. coli.     

Indole-3-acetic acid in the culture medium of two axenic green microalgae

In the view of the facts that algal extracts have been used in agriculture asa source of plant growth stimulating agents and IAA has been shown to bepresent in the extracts, a study was planned to establish whether or notaxenic algae can produce IAA. Evidence is provided for extracellular IAAproduction during culture of two axenic green microalgae. IAAidentification was based on co-chromatography with the standard, analysisof UV and fluorescent spectra, and gas chromatography – selectedion-monitoring mass spectrometry. HPLC analyses showed that underthe experimental conditions the amounts of IAA released to the mediumby Scenedesmus armatus and Chlorella pyrenoidosa weregenerally low. IAA tended to occur in Scenedesmus armatus culturemedium at higher concentrations than in that of Chlorellapyrenoidosa. In fast-growing cultures of Scenedesmus armatus,constantly aerated with CO₂/air mixture, the concentration of IAAcalculated per cell was less than in the slow-growing cultures.     

Endothelial heparan sulphate: Compositional analysis and comparison of chains from different proteoglycan populations

From cultures of human umbilical vein endothelial cells incubated with³H-glucosamine or³⁵S-sulphate, we have purified three heparan sulphate proteoglycans: 1) a low density (1.31 g/ml) proteoglycan from the cell extract, 2) a low density proteoglycan from the medium, and 3) a high density (>1.4 g/ml) proteoglycan from the medium. The disaccharide composition of heparan sulphate chains from the low density proteoglycan of the medium was examined, using specific chemical and enzymic degradations followed by gel chromatography and strong anion exchange HPLC. Chains released from each of the different proteoglycan populations were then compared by gel chromatography and gradient polyacrylamide gel electrophoresis before and after various specific degradations. The results indicate that heparan sulphate from human endothelial cells are large polymers (MW>50,000) of low overall sulphation (32–35%N-sulphated glucosamine and an N/O-linked sulphate ratio of 2.0) with rare and solitary heparin-like disaccharides. Heparan sulphate from the different proteoglycan populations appeared to have similar structure except that chains from the high density fraction were larger polymers.Abbreviations HSPG heparan sulphate proteoglycan - DSPG dermatan sulphate proteoglycan - GlcNAc(6S) N-acetylglucosamine 6-sulphate - GlcNAc6R glucosamine with either-OH or-OSO₃ at C-6 - GlcNR glucosamine with either-SO₃ or-COCH₃ as N-substituent - GlcNSO₃ N-sulphated glucosamine - GlcNSO₃(3S) N-sulphated glucosamine 3-sulphate - GlcA d-glucuronic acid - IdoA l-iduronic acid - IdoA(2S) iduronic acid 2-sulphate - HexA hexuronic acid - DHexA hexuronic acid with a 4,5-double bond - Xyl xylose - SAX strong anion exchange - d.p. degree of polymerization (a disaccharide has d.p.=1 etc) - AUFS absorbance units full scale The codes used for proteoglycans denote in turn: C 2, low-density (1.35–1.28 g/ml) HSPG from the cell extract; M 1a, high density (>1.4 g/ml) HSPG fraction from the spent medium; M 2a, low-density (1.31 g/ml) HSPG from the spent medium [6].     

Photochemical activities,pigment distribution and photosynthetic unit size of subchloroplast particles isolated from synchronized cells of Scenedesmus obliquus

Photosystem II (PS II) reactions of chloroplast particles show the same variations during the synchronous life cycle of Scenedesmus obliquus, strain D₃ (Gaffron Biol. Zbl. 59, 302 1939), as the whole cells they derived from. Photosystem I (PS I) reactions of whole cells and of subchloroplast particles show little or no variation in their activity, whereas PS I reactions of chloroplast particles vary like PS II reactions during the life cycle. The variation in chloroplast particles could be attributed to the change in the reoxidation capacity of plastoquinone still attached to PS I. Digitonin-treatment of chloroplast particles from Scenedesmus and subsequent sucrose density gradient separation yielded 3 distinct fractions: Fraction I contained pure PS I particles with the most efficient PS I-mediated methylviologen (MV) reduction with subsequent oxygen uptake (3 mmol O₂/mg Chl·h); no Hill reaction; and a high chlorophyll a/b ratio, and a vast amount of unbound protein xanthophyll complexes. Fraction II is enriched in PS II particles, with little PS I activity (less than 10% of the PS I particles) and a low chlorophyll a/b ratio. The activity of the water-splitting system was completely lost. This fraction must also contain most of the light-harvesting pigment system. Fraction III is also enriched in PS II with even less PS I activity, but the ratio of chlorophyll a/b is slightly higher than in whole cells and the water-splitting system is intact. -carotene was part of all fractions whereas functional xanthophylls seemed to be restricted to the PS II particles. From the constant chlorophyll P/700 ratio we had to conclude that size of the photosynthetic unit does not change during the life cycle of a synchronized Scenedesmus obliquus culture.Abbreviations DBMIB 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethyl-urea - DCPIP dichlorphenolindophenol - MV methylviologen (paraquat) - PS I photosystem I - PS II photosystem II - DPC diphenyl-carbazide     

Isolation and characterization of sulfur globule proteins from Chromatium vinosum and Thiocapsa roseopersicina

Purple sulfur bacteria store sulfur as intracellular globules enclosed by a protein envelope. The proteins associated with sulfur globules of Chromatium vinosum and Thiocapsa roseopersicina were isolated by extraction into 50% aqueous acetonitrile containing 1% trifluoroacetic acid and 10 mM dithiothreitol. The extracted proteins were separated by reversed-phase HPLC, revealing three major proteins from C. vinosum and two from T. roseopersicina. All of these proteins have similar, rather unusual amino acid compositions, being rich in glycine and aromatic amino acids, particularly tyrosine. The molecular masses of the C. vinosum proteins were determined to be 10,498, 10,651, and 8,479 Da, while those from T. roseopersicina were found to be 10,661 and 8,759 Da by laser desorption time-of-flight mass spectrometry. The larger T. roseopersicina protein is N-terminally blocked, probably by acetylation, but small amounts of the unblocked form (mass = 10,619) were also isolated by HPLC. Protein sequencing showed that the two larger C. vinosum proteins are homologous to each other and to the large T. roseopersicina protein. The 8,479 Da C. vinosum and 8,759 Da T. roseopersicina proteins are also homologous, indicating that sulfur globule proteins are conserved between different species of purple sulfur bacteria.Abbreviations BNPS-skatole 2 (2-Nitrophenylsulfenyl)-3-methyl-3-bromoindolenine - CNB Cyanogen bromide - Cv1, Cv2, and Cv3 Chromatium vinosum sulfur globule proteins - SGP and SGPs Sulfur globule protein(s) - TFA Trifluoroacetic acid - Tr0, Tr1, and Tr2 Thiocapsa roseopersicina sulfur globule proteins