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The salt-tolerant marine microalgae Dunaliella tertiolecta is reported to generate significant amounts of intracellular glycerol as an osmoprotectant under high salt conditions. This study highlights the phylogenetic distribution and comparative glycerol biosynthesis of seven new Dunaliella isolates compared to a D. tertiolecta reference strain. Phylogenetic analysis indicates that all Dunaliella isolates are newly discovered and do not relate to the D. tertiolecta reference. Several studies have identified light color and intensity and salt concentration alone as the most inducing factors impacting glycerol productivity. This study aims to optimize glycerol production by investigating these described factors singularly and in combination to improve the glycerol product titer. Glycerol production data indicate that cultivation with white light of an intensity between 500 and 2000 µmol m-2 s-1 as opposed to 100 µmol m-2 s-1 achieves higher biomass and thereby higher glycerol titers for all our tested Dunaliella strains. Moreover, applying higher light intensity in a cultivation of 1.5 M NaCl and an increase to 3 M NaCl resulted in hyperosmotic stress conditions, providing the highest glycerol titer. Under these optimal light intensity and salt conditions, the glycerol titer of D. tertiolecta could be doubled to 0.79 mg mL-1 in comparison to 100 µmol m-2 s-1 and salt stress to 2 M NaCl, and was higher compared to singularly optimized conditions. Furthermore, under the same conditions, glycerol extracts from new Dunaliella isolates did provide up to 0.94 mg mL-1. This highly pure algae-glycerol obtained under optimal production conditions can find widespread applications, e.g., in the pharmaceutical industry or the production of sustainable carbon fibers.
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Lipopeptides are surface active, natural products of bacteria, fungi and green-blue algae origin, having diverse structures and functionalities. In analogy, a number of chemical synthesis techniques generated new designer lipopeptides with desirable features and functions. Lipopetides are self-assembly guided, supramolecular compounds which have the capacity of high-density presentation of the functional epitopes at the surface of the nanostructures. This feature contributes to their successful application in several industry sectors, including food, feed, personal care, and pharmaceutics. In this comprehensive review, the novel class of ribosomally synthesized lipopeptides is introduced alongside the more commonly occuring non-ribosomal lipopeptides. We highlight key representatives of the most researched as well as recently described lipopeptide families, with emphasis on structural features, self-assembly and associated functions. The common biological, chemical and hybrid production routes of lipopeptides, including prominent analogues and derivatives are also discussed. Furthermore, genetic engineering strategies aimed at increasing lipopeptide yields, diversity and biological activity are summarized and exemplified. With respect to application, this work mainly details the potential of lipopeptides in personal care and cosmetics industry as cleansing agents, moisturizer, anti-aging/anti-wrinkling, skin whitening and preservative agents as well as the pharmaceutical industry as anitimicrobial agents, vaccines, immunotherapy, and cancer drugs. Given that this review addresses human applications, we conclude on the topic of safety of lipopeptide formulations and their sustainable production.
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Cosméticos , Lipopéptidos , Humanos , Lipopéptidos/química , Bacterias , Ingeniería Genética , Cosméticos/química , Preparaciones Farmacéuticas , Tensoactivos/químicaRESUMEN
The potential of fungi for use as biotechnological factories in the production of a range of valuable metabolites, such as enzymes, terpenes, and volatile aroma compounds, is high. Unlike other microorganisms, fungi mostly secrete secondary metabolites into the culture medium, allowing for easy extraction and analysis. To date, the most commonly used technique in the analysis of volatile organic compounds (VOCs) is gas chromatography, which is time and labour consuming. We propose an alternative ambient screening method that provides rapid chemical information for characterising the VOCs of filamentous fungi in liquid culture using a commercially available ambient dielectric barrier discharge ionisation (DBDI) source connected to a quadrupole-Orbitrap mass spectrometer. The effects of method parameters on measured peak intensities of a series of 8 selected aroma standards were optimised with the best conditions being selected for sample analysis. The developed method was then deployed to the screening of VOCs from samples of 13 fungal strains in three different types of complex growth media showing clear differences in VOC profiles across the different media, enabling determination of best culturing conditions for each compound-strain combination. Our findings underline the applicability of ambient DBDI for the direct detection and comparison of aroma compounds produced by filamentous fungi in liquid culture.
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Compuestos Orgánicos Volátiles , Compuestos Orgánicos Volátiles/análisis , Cromatografía de Gases y Espectrometría de Masas/métodos , Espectrometría de Masas , Medios de Cultivo/análisis , HongosRESUMEN
Due to increasing oil prices and climate change concerns, biofuels have become increasingly important as potential alternative energy sources. However, the use of arable lands and valuable resources for the production of biofuel feedstock compromises food security and negatively affect the environment. Single cell oils (SCOs), accumulated by oleaginous yeasts, show great promise for efficient production of biofuels. However, the high production costs attributed to feedstocks or raw materials present a major limiting factor. The fermentative conversion of abundant, low-value biomass into microbial oil would alleviate this limitation. Here, we explore the feasibility of utilizing microalgae-based cell residues as feedstock for yeast oil production. We developed an efficient, single-step enzymatic hydrolysis to generate Scenedesmus obtusiusculus hydrolysate (SH) without thermo-chemical pretreatment. With this eco-friendly process, glucose conversion efficiencies reached 90-100%. Cutaneotrichosporon oleaginosus, Cryptococcus curvatus and Rhodosporidium toruloides were cultivated on SH as sole nutrients source. Only C. oleaginosus was able to accumulate intracellular lipids, with a 35% (g lipid/g DCW) content and a yield of 3.6 g/L. Our results demonstrate the potential valorization of algal biomass into desired end-products such as biofuels.
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Basidiomycota/crecimiento & desarrollo , Biomasa , Lípidos/biosíntesis , Microalgas/química , Rhodotorula/crecimiento & desarrollo , Scenedesmus/químicaRESUMEN
BACKGROUND: Oleaginous organisms are a promising, renewable source of single cell oil. Lipid accumulation is mainly induced by limitation of nutrients such as nitrogen, phosphorus or sulfur. The oleaginous yeast Trichosporon oleaginosus accumulates up to 70% w/w lipid under nitrogen stress, while cultivation in non-limiting media only yields 9% w/w lipid. Uncoupling growth from lipid accumulation is key for the industrial process applicability of oleaginous yeasts. This study evaluates the effects of rapamycin on TOR specific signaling pathways associated with lipogenesis in Trichosporon oleaginosus for the first time. RESULTS: Supplementation of rapamycin to nutrient rich cultivation medium led to an increase in lipid yield of up to 38% g/L. This effect plateaued at 40 µM rapamycin. Interestingly, the fatty acid spectrum resembled that observed with cultivation under nitrogen limitation. Significant changes in growth characteristics included a 19% increase in maximum cell density and a 12% higher maximum growth rate. T. oleaginosus only has one Tor gene much like the oleaginous yeast Rhodosporidium toruloides. Consequently, we analyzed the effect of rapamycin on T. oleaginosus specific TORC signaling using bioinformatic methodologies. CONCLUSIONS: We confirm, that target of rapamycin complex 1 (TORC1) is involved in control of lipid production and cell proliferation in T. oleaginosus and present a homology based signaling network. Signaling of lipid induction by TORC1 and response to carbon depletion to this complex appear to be conserved, whereas response to nitrogen limitation and autophagy are not. This work serves as a basis for further investigation regarding the control and induction of lipid accumulation in oil yeasts.
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Proliferación Celular/fisiología , Lipogénesis/fisiología , Complejos Multiproteicos/metabolismo , Aceites/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Trichosporon/fisiología , Diana Mecanicista del Complejo 1 de la Rapamicina , Especificidad de la Especie , Trichosporon/clasificaciónRESUMEN
Microalgae are capable of producing up to 70% w/w triglycerides with respect to their dry cell weight. Since microalgae utilize the greenhouse gas CO2, they can be cultivated on marginal lands and grow up to ten times faster than terrestrial plants, the generation of algae oils is a promising option for the development of sustainable bioprocesses, that are of interest for the chemical lubricant, cosmetic and food industry. For the first time we have carried out the optimization of supercritical carbon dioxide (SCCO2) mediated lipid extraction from biomass of the microalgae Scenedesmus obliquus and Scenedesmus obtusiusculus under industrrially relevant conditions. All experiments were carried out in an industrial pilot plant setting, according to current ATEX directives, with batch sizes up to 1.3 kg. Different combinations of pressure (7-80 MPa), temperature (20-200 °C) and CO2 to biomass ratio (20-200) have been tested on the dried biomass. The most efficient conditions were found to be 12 MPa pressure, a temperature of 20 °C and a CO2 to biomass ratio of 100, resulting in a high extraction efficiency of up to 92%. Since the optimized CO2 extraction still yields a crude triglyceride product that contains various algae derived contaminants, such as chlorophyll and carotenoids, a very effective and scalable purification procedure, based on cost efficient bentonite based adsorbers, was devised. In addition to the sequential extraction and purification procedure, we present a consolidated online-bleaching procedure for algae derived oils that is realized within the supercritical CO2 extraction plant.
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Microalgas , Biomasa , Dióxido de Carbono , Lípidos , ScenedesmusRESUMEN
Isobutanol is deemed to be a next-generation biofuel and a renewable platform chemical.1 Non-natural biosynthetic pathways for isobutanol production have been implemented in cell-based and in vitro systems with Bacillus subtilis acetolactate synthase (AlsS) as key biocatalyst.2-6 AlsS catalyzes the condensation of two pyruvate molecules to acetolactate with thiamine diphosphate and Mg(2+) as cofactors. AlsS also catalyzes the conversion of 2-ketoisovalerate into isobutyraldehyde, the immediate precursor of isobutanol. Our phylogenetic analysis suggests that the ALS enzyme family forms a distinct subgroup of ThDP-dependent enzymes. To unravel catalytically relevant structure-function relationships, we solved the AlsS crystal structure at 2.3 Å in the presence of ThDP, Mg(2+) and in a transition state with a 2-lactyl moiety bound to ThDP. We supplemented our structural data by point mutations in the active site to identify catalytically important residues.
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Acetolactato Sintasa/química , Bacillus subtilis/química , Proteínas Bacterianas/química , Butanoles/química , Acetolactato Sintasa/genética , Acetolactato Sintasa/metabolismo , Aldehídos/química , Aldehídos/metabolismo , Bacillus subtilis/clasificación , Bacillus subtilis/enzimología , Bacillus subtilis/genética , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Biocatálisis , Biocombustibles , Butanoles/metabolismo , Dominio Catalítico , Cationes Bivalentes , Coenzimas/química , Coenzimas/metabolismo , Cristalografía por Rayos X , Escherichia coli/genética , Escherichia coli/metabolismo , Expresión Génica , Hemiterpenos , Cetoácidos/química , Cetoácidos/metabolismo , Lactatos/química , Lactatos/metabolismo , Magnesio/química , Magnesio/metabolismo , Modelos Moleculares , Filogenia , Mutación Puntual , Ácido Pirúvico/química , Ácido Pirúvico/metabolismo , Relación Estructura-Actividad , Tiamina Pirofosfato/química , Tiamina Pirofosfato/metabolismoRESUMEN
There is a considerable potential of cold-active biocatalysts for versatile industrial applications. A psychrophilic bacterial strain, Shewanella arctica 40-3, has been isolated from arctic sea ice and was shown to exhibit pullulan-degrading activity. Purification of a monomeric, 150-kDa pullulanase was achieved using a five-step purification approach. The native enzyme was purified 50.0-fold to a final specific activity of 3.0 U/mg. The enzyme was active at a broad range of temperature (10-50 °C) and pH (5-9). Optimal activity was determined at 45 °C and pH 7. The presence of various metal ions is tolerated by the pullulanase, while detergents resulted in decreased activity. Complete conversion of pullulan to maltotriose as the sole product and N-terminal amino acid sequence indicated that the enzyme is a type-I pullulanase and belongs to rarely characterized pullulan-degrading enzymes from psychrophiles.
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Proteínas Bacterianas/química , Frío , Glicósido Hidrolasas/química , Shewanella/enzimología , Adaptación Fisiológica , Secuencia de Aminoácidos , Regiones Árticas , Proteínas Bacterianas/metabolismo , Glicósido Hidrolasas/metabolismo , Datos de Secuencia MolecularRESUMEN
A novel aerobic, psychrotolerant marine bacterium was isolated at 4 degrees C from seawater samples collected from Spitzbergen in the Arctic. The strain was a polar-flagellated, Gram-negative bacterium that grew optimally at 10-15 degrees C and pH 7-8 in media containing 2-3% NaCl (w/v), using various carbohydrates and organic acids as substrates. The main fatty acid components included 16:0 (12.7% of total fatty acids), straight-chain saturated fatty acid methyl ester (FAME) and 16:1omega7c (40.2%) monounsaturated FAME. Phylogenetic analysis revealed a close relationship (99% 16S rRNA gene sequence similarity) between the novel isolate and Pseudoalteromonas elyakovii KMM 162T and some other species of the genus Pseudoalteromonas. The DNA G+C content of the novel strain was 39 mol%. DNA-DNA hybridization showed only 47.6% DNA-DNA relatedness with P. elyakovii KMM 162T, 44.2% with Pseudoalteromonas distincta KMM 638T and 22.6% with Pseudoalteromonas nigrifaciens NCIMB 8614T. Based on phylogenetic and phenotypic characteristics, this isolate represents a novel species of the genus Pseudoalteromonas for which the name Pseudoalteromonas arctica is proposed; the type strain is A 37-1-2T (=LMG 23753T=DSM 18437T).