Heuristic Modeling and 3D Stereoscopic Visualization of a Chlamydomonas reinhardtii Cell.

The structural modeling and representation of cells is a complex task as different microscopic, spectroscopic and other information resources have to be combined to achieve a three-dimensional representation with high accuracy. Moreover, to provide an appropriate spatial representation of the cell, a stereoscopic 3D (S3D) visualization is favorable. In this work, a structural cell model is created by combining information from various light microscopic and electron microscopic images as well as from publication-related data. At the mesoscopic level each cell component is presented with special structural and visual properties; at the molecular level a cell membrane composition and the underlying modeling method are discussed; and structural information is correlated with those at the functional level (represented by simplified energy-producing metabolic pathways). The organism used as an example is the unicellular Chlamydomonas reinhardtii, which might be important in future alternative energy production processes. Based on the 3D model, an educative S3D animation was created which was shown at conferences. The complete workflow was accomplished by using the open source 3D modeling software Blender. The discussed project including the animation is available from: http://Cm5.CELLmicrocosmos.org.

BiPACE 2D--graph-based multiple alignment for comprehensive 2D gas chromatography-mass spectrometry.

MOTIVATION: Comprehensive 2D gas chromatography-mass spectrometry is an established method for the analysis of complex mixtures in analytical chemistry and metabolomics. It produces large amounts of data that require semiautomatic, but preferably automatic handling. This involves the location of significant signals (peaks) and their matching and alignment across different measurements. To date, there exist only a few openly available algorithms for the retention time alignment of peaks originating from such experiments that scale well with increasing sample and peak numbers, while providing reliable alignment results. RESULTS: We describe BiPACE 2D, an automated algorithm for retention time alignment of peaks from 2D gas chromatography-mass spectrometry experiments and evaluate it on three previously published datasets against the mSPA, SWPA and Guineu algorithms. We also provide a fourth dataset from an experiment studying the H2 production of two different strains of Chlamydomonas reinhardtii that is available from the MetaboLights database together with the experimental protocol, peak-detection results and manually curated multiple peak alignment for future comparability with newly developed algorithms. AVAILABILITY AND IMPLEMENTATION: BiPACE 2D is contained in the freely available Maltcms framework, version 1.3, hosted at http://maltcms.sf.net, under the terms of the L-GPL v3 or Eclipse Open Source licenses. The software used for the evaluation along with the underlying datasets is available at the same location. The C.reinhardtii dataset is freely available at http://www.ebi.ac.uk/metabolights/MTBLS37.

Synthesis of transparent aminosilane-derived silica based networks for entrapment of sensitive materials.

A novel sol-gel synthesis route is reported which results in the formation of optically transparent silica based hydro- and xerogels from an aminosilane precursor in aqueous solutions. These materials can be used for entrapment of microalgae and light-harvesting complex (LHC) samples.

Identification of Monoraphidium contortum as a promising species for liquid biofuel production.

In this work, 30 microalgae strains from 17 genera were investigated in regard to biomass productivity in photoautotrophic growth conditions, lipid amount, lipid quality and biomass degradability. Six strains could be identified with robust phototrophic growth properties and high biomass productivities equal or above 300 mg l(-1) day(-1). Anaerobic fermentation of the algal biomass was most efficient for the marine members of the genera Dunaliella and Navicula, while biogas production with the freshwater strains generally resulted in lower methane yields. Monoraphidium contortum was identified as promising candidate for liquid biofuel production, characterized by high biomass productivity during maximum growth (maximum increase of 896 mg dry biomass weight (DW) l(-1) day(-1)) and a promising lipid profile. Neutral lipid production was strongly induced in M. contortum by nitrogen deficient conditions and accumulated to up to 20.4±2.2% of DW.

Cellulose degradation and assimilation by the unicellular phototrophic eukaryote Chlamydomonas reinhardtii.

Plants convert sunlight to biomass, which is primarily composed of lignocellulose, the most abundant natural biopolymer and a potential feedstock for fuel and chemical production. Cellulose assimilation has so far only been described for heterotrophic organisms that rely on photosynthetically active primary producers of organic compounds. Among phototrophs, the unicellular green microalga Chlamydomonas reinhardtii is widely known as one of the best established model organisms. It occupies many habitats, including aquatic and soil ecosystems. This ubiquity underscores the versatile metabolic properties of this microorganism. Here we present yet another paradigm of adaptation for C. reinhardtii, highlighting its photoheterotrophic ability to utilize cellulose for growth in the absence of other carbon sources. When grown under CO(2)-limiting conditions in the light, secretion of endo-ß-1,4-glucanases by the cell causes digestion of exogenous cellulose, followed by cellobiose uptake and assimilation. Phototrophic microbes like C. reinhardtii may thus serve as biocatalysts for cellulosic biofuel production.

Time-course global expression profiles of Chlamydomonas reinhardtii during photo-biological H2 production.

We used a microarray study in order to compare the time course expression profiles of two Chlamydomonas reinhardtii strains, namely the high H2 producing mutant stm6glc4 and its parental WT strain during H2 production induced by sulfur starvation. Major cellular reorganizations in photosynthetic apparatus, sulfur and carbon metabolism upon H2 production were confirmed as common to both strains. More importantly, our results pointed out factors which lead to the higher H2 production in the mutant including a higher starch accumulation in the aerobic phase and a lower competition between the H2ase pathway and alternative electron sinks within the H2 production phase. Key candidate genes of interest with differential expression pattern include LHCSR3, essential for efficient energy quenching (qE). The reduced LHCSR3 protein expression in mutant stm6glc4 could be closely related to the high-light sensitive phenotype. H2 measurements carried out with the LHCSR3 knock-out mutant npq4 however clearly demonstrated that a complete loss of this protein has almost no impact on H2 yields under moderate light conditions. The nuclear gene disrupted in the high H2 producing mutant stm6glc4 encodes for the mitochondrial transcription termination factor (mTERF) MOC1, whose expression strongly increases during -S-induced H2 production in WT strains. Studies under phototrophic high-light conditions demonstrated that the presence of functional MOC1 is a prerequisite for proper LHCSR3 expression. Furthermore knock-down of MOC1 in a WT strain was shown to improve the total H2 yield significantly suggesting that this strategy could be applied to further enhance H2 production in other strains already displaying a high H2 production capacity. By combining our array data with previously published metabolomics data we can now explain some of the phenotypic characteristics which lead to an elevated H2 production in stm6glc4.

The interplay of proton, electron, and metabolite supply for photosynthetic H2 production in Chlamydomonas reinhardtii.

To obtain a detailed picture of sulfur deprivation-induced H(2) production in microalgae, metabolome analyses were performed during key time points of the anaerobic H(2) production process of Chlamydomonas reinhardtii. Analyses were performed using gas chromatography coupled to mass spectrometry (GC/MS), two-dimensional gas chromatography combined with time-of-flight mass spectrometry (GCxGC-TOFMS), lipid and starch analysis, and enzymatic determination of fermentative products. The studies were designed to provide a detailed metabolite profile of the solar Bio-H(2) production process. This work reports on the differential analysis of metabolic profiles of the high H(2)-producing strain Stm6Glc4 and the wild-type cc406 (WT) before and during the H(2) production phase. Using GCxGC-TOFMS analysis the number of detected peaks increased from 128 peaks, previously detected by GC/MS techniques, to ∼1168. More detailed analysis of the anaerobic H(2) production phase revealed remarkable differences between wild-type and mutant cells in a number of metabolic pathways. Under these physiological conditions the WT produced up to 2.6 times more fatty acids, 2.2 times more neutral lipids, and up to 4 times more fermentation products compared with Stm6Glc4. Based on these results, specific metabolic pathways involving the synthesis of fatty acids, neutral lipids, and fermentation products during anaerobiosis in C. reinhardtii have been identified as potential targets for metabolic engineering to further enhance substrate supply for the hydrogenase(s) in the chloroplast.

The metabolome of Chlamydomonas reinhardtii following induction of anaerobic H2 production by sulfur depletion.

The metabolome of the model species Chlamydomonas reinhardtii has been analyzed during 120 h of sulfur depletion to induce anaerobic hydrogen (H(2)) production, using NMR spectroscopy, gas chromatography coupled to mass spectrometry, and TLC. The results indicate that these unicellular green algae consume freshly supplied acetate in the medium to accumulate energy reserves during the first 24 h of sulfur depletion. In addition to the previously reported accumulation of starch, large amounts of triacylglycerides were deposited in the cells. During the early 24- to 72-h time period fermentative energy metabolism lowered the pH, H(2) was produced, and amino acid levels generally increased. In the final phase from 72 to 120 h, metabolism slowed down leading to a stabilization of pH, even though some starch and most triacylglycerides remained. We conclude that H(2) production does not slow down due to depletion of energy reserves but rather due to loss of essential functions resulting from sulfur depletion or due to a build-up of the toxic fermentative products formate and ethanol.

Functional integration of the HUP1 hexose symporter gene into the genome of C. reinhardtii: Impacts on biological H(2) production.

Phototrophic organisms use photosynthesis to convert solar energy into chemical energy. In nature, the chemical energy is stored in a diverse range of biopolymers. These sunlight-derived, energy-rich biopolymers can be converted into environmentally clean and CO(2) neutral fuels. A select group of photosynthetic microorganisms have developed the ability to extract and divert protons and electrons derived from water to chloroplast hydrogenase(s) to produce molecular H(2) fuel. Here, we describe the development and characterization of C. reinhardtii strains, derived from the high H(2) production mutant Stm6, into which the HUP1 (hexose uptake protein) hexose symporter from Chlorella kessleri was introduced. The isolated cell lines can use externally supplied glucose for heterotrophic growth in the dark. More importantly, external glucose supply (1mM) was shown to increase the H(2) production capacity in strain Stm6Glc4 to approximately 150% of that of the high-H(2) producing strain, Stm6. This establishes the foundations for a new fuel production process in which H(2)O and glucose can simultaneously be used for H(2) production. It also opens new perspectives on future strategies for improving bio-H(2) production efficiency under natural day/night regimes and for using sugar waste material for energy production in green algae as photosynthetic catalysts.