RESUMEN
In recent years, bio-based production of free fatty acids from renewable resources has attracted attention for their potential as precursors for the production of biofuels and biochemicals. In this study, the oleaginous yeast Yarrowia lipolytica was engineered to produce free fatty acids by eliminating glycerol metabolism. Free fatty acid production was monitored under lipogenic conditions with glycerol as a limiting factor. Firstly, the strain W29 (Δgpd1), which is deficient in glycerol synthesis, was obtained. However, W29 (Δgpd1) showed decreased biomass accumulation and glucose consumption in lipogenic medium containing a limiting supply of glycerol. Analysis of substrate utilization from a mixture of glucose and glycerol by the parental strain W29 revealed that glycerol was metabolized first and glucose utilization was suppressed. Thus, the Δgpd1Δgut2 double mutant, which is deficient also in glycerol catabolism, was constructed. In this genetic background, growth was repressed by glycerol. Oleate toxicity was observed in the Δgpd1Δgut2Δpex10 triple mutant strain which is deficient additionally in peroxisome biogenesis. Consequently, two consecutive rounds of selection of spontaneous mutants were performed. A mutant released from growth repression by glycerol was able to produce 136.8 mg L-1 of free fatty acids in a test tube, whereas the wild type accumulated only 30.2 mg L-1 . Next, an isolated oleate-resistant strain produced 382.8 mg L-1 of free fatty acids. Finely, acyl-CoA carboxylase gene (ACC1) over-expression resulted to production of 1436.7 mg L-1 of free fatty acids. The addition of dodecane promoted free fatty acid secretion and enhanced the level of free fatty acids up to 2033.8 mg L-1 during test tube cultivation.
Asunto(s)
Ácidos Grasos no Esterificados/metabolismo , Glicerol/metabolismo , Ingeniería Metabólica/métodos , Yarrowia/metabolismo , Proliferación Celular/efectos de los fármacos , Ácidos Grasos no Esterificados/análisis , Ácidos Grasos no Esterificados/toxicidad , Glucosa/metabolismo , Glicerol/farmacología , Redes y Vías Metabólicas/efectos de los fármacos , Redes y Vías Metabólicas/genética , Ácido Oléico/metabolismo , Ácido Oléico/toxicidad , Yarrowia/efectos de los fármacos , Yarrowia/genéticaRESUMEN
Bio-based succinic acid production can redirect industrial chemistry processes from using limited hydrocarbons to renewable carbohydrates. A fermentation process that does not require pH-titrating agents will be advantageous to the industry. Previously, a Yarrowia lipolytica strain that was defective for succinate dehydrogenase was constructed and was found to accumulate up to 17.5 g L(-1) of succinic acid when grown on glycerol without buffering. Here, a derivative mutant was isolated that produced 40.5 g L(-1) of succinic acid in 36 h with a yield of 0.32 g g(-1) glycerol. A combination approach of induced mutagenesis and metabolic evolution allowed isolation of another derivative that could utilize glucose efficiently and accumulated 50.2 g L(-1) succinic acid in 54 h with a yield of 0.43 g g(-1) . The parent strain of these isolated mutants was used for [1,6-(13) C2 ]glucose assimilation analysis. At least 35% glucose was estimated to be utilized through the pentose phosphate pathway, while ≥84% succinic acid was formed through the oxidative branch of the tricarboxylic acid cycle. Biotechnol. Bioeng. 2016;113: 2425-2432. © 2016 Wiley Periodicals, Inc.
Asunto(s)
Isótopos de Carbono/farmacocinética , Análisis de Flujos Metabólicos/métodos , Succinato Deshidrogenasa/metabolismo , Ácido Succínico/metabolismo , Yarrowia/fisiología , Glucosa/metabolismo , Tasa de Depuración Metabólica , Succinato Deshidrogenasa/deficiencia , Succinato Deshidrogenasa/genéticaRESUMEN
The atomic structure and elastic properties of Y-shaped silicon nanowires of "fork"- and "bough"-types were theoretically studied, and effective Young moduli were calculated using Tersoff interatomic potential. The oscillation of fork Y-type branched nanowires with various branch lengths and diameters was studied. In the final stages of the bending, the formation of new bonds between different parts of the wires was observed. It was found that the stiffness of the nanowires is comparable with the stiffness of Y-shaped carbon nanotubes.
Asunto(s)
Elasticidad , Modelos Moleculares , Nanocables/química , Silicio/química , Módulo de Elasticidad , Conformación Molecular , Estrés MecánicoRESUMEN
Sulfur deprivation of algal cultures selectively and partially inactivates photosystem II (PSII)-catalyzed O(2) evolution, induces anaerobiosis and hydrogenase expression, and results in sustained H(2) photoproduction for several days. We show that re-addition of limiting amounts of sulfate (1-10 microM final concentration) to the cultures during the H(2)-production phase temporarily reactivates PSII photochemical and O(2)-evolution activity and re-establishes higher rates of electron transport through the photosynthetic electron transport chain. The reactivation of PSII occurs by de novo D1 protein synthesis, but does not result in the re-establishment of aerobic conditions in the reactor, detectable by dissolved-O(2) sensors. However, concomitant H(2) photoproduction is inhibited, possibly due to excessive intra-cellular levels of photosynthetically-evolved O(2). The partial recovery of electron transport rates correlates with the re-oxidation of the plastoquinone (PQ) pool, as observed by pulse-amplitude modulated (PAM) and fluorescence-induction measurements. These results show that the presence of a more oxidized PQ pool releases some of the down-regulation of electron transport caused by the anaerobic conditions.
Asunto(s)
Chlamydomonas reinhardtii/efectos de los fármacos , Chlamydomonas reinhardtii/metabolismo , Hidrógeno/metabolismo , Sulfatos/farmacología , Azufre/deficiencia , Animales , Transporte de Electrón/efectos de los fármacos , Fluorescencia , Oxígeno/metabolismo , Fotosíntesis/efectos de los fármacos , Fotosíntesis/fisiología , Plastoquinona/metabolismo , Sulfatos/metabolismo , Azufre/metabolismo , Factores de TiempoRESUMEN
This study demonstrates, for the first time, that it is possible to couple sulfate-limited Chlamydomonas reinhardtii growth to continuous H2 photoproduction for more than 4000 h. A two-stage chemostat system physically separates photosynthetic growth from H2 production, and it incorporates two automated photobioreactors (PhBRs). In the first PhBR, the algal cultures are grown aerobically in chemostat mode under limited sulfate to obtain photosynthetically competent cells. Active cells are then continuously delivered to the second PhBR, where H2 production occurs under anaerobic conditions. The dependence of the H2 production rate on sulfate concentration in the medium, dilution rates in the PhBRs, and incident light intensity is reported.