SEARCH · Engineering Papers
Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.
Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.
Cell walls in grasses contain arabinoxylan polysaccharides decorated with ferulate groups but the mechanism by which the ferulate is attached to arabinoxylans has long remained unknown. Here, a new study shows that ferulate is transferred to arabinoxylan from a 3,6-di-O-feruloyl sucrose intermediate formed in a sucrose ferulate cycle.
Abstract Engineered cyanobacterium Synechococcus elongatus can use light and CO 2 to produce sucrose, making it a promising candidate for use in co-cultures with heterotrophic workhorses. However, this process is challenged by the mutual stresses generated from the multispecies microbial culture. Here we demonstrate an ecosystem where S. elongatus is freely grown in a photo-bioreactor (PBR) containing an engineered heterotrophic workhorse (either β-carotene-producing Yarrowia lipolytica or indigoidine-producing Pseudomonas putida ) encapsulated in calcium-alginate hydrogel beads. The encapsulation prevents growth interference, allowing the cyanobacterial culture to produce high sucrose concentrations enabling the production of indigoidine and β-carotene in the heterotroph. Our experimental PBRs yielded an indigoidine titer of 7.5 g/L hydrogel and a β-carotene titer of 1.3 g/L hydrogel, amounts 15–22-fold higher than in a comparable co-culture without encapsulation. Moreover, 13 C-metabolite analysis and protein overexpression tests indicated that the hydrogel beads provided a favorable microenvironment where the cell metabolism inside the hydrogel was comparable to that in a free culture. Finally, the heterotroph-containing hydrogels were easily harvested and dissolved by EDTA for product recovery, while the cyanobacterial culture itself could be reused for the next batch of immobilized heterotrophs. This co-cultivation and hydrogel encapsulation system is a successful demonstration of bioprocess optimization under photobioreactor conditions.
Abstract Metabolic engineering to divert carbon flux from sucrose to oil in high biomass crop like sugarcane is an emerging strategy to boost lipid yields per hectare for biodiesel production. Sugarcane stems comprise more than 70% of the crops' biomass and can accumulate sucrose in excess of 20% of their extracted juice. The energy content of oils in the form of triacylglycerol (TAG) is more than twofold that of carbohydrates. Here, we report a step change in TAG accumulation in sugarcane stem tissues achieving an average of 4.3% of their dry weight (DW) in replicated greenhouse experiments by multigene engineering. The metabolic engineering included constitutive co‐expression of wrinkled1; diacylglycerol acyltransferase1‐2; cysteine‐oleosin; and ribonucleic acid interference‐suppression of sugar‐dependent1. The TAG content in leaf tissue was also elevated by more than 400‐fold compared to non‐engineered sugarcane to an average of 8.0% of the DW and the amount of total fatty acids reached about 13% of the DW. With increasing TAG accumulation an increase of 18:1 unsaturated fatty acids was observed at the expense of 16:0 and 18:0 saturated fatty acids. Total biomass accumulation, soluble lignin, Brix and juice content were significantly reduced in the TAG hyperaccumulating sugarcane lines. Overcoming this yield drag by engineering lipid accumulation into late stem development will be critical to exceed lipid yields of current oilseed crops.
Abstract Background The industrial feasibility of photosynthetic bioproduction using cyanobacterial platforms remains challenging due to insufficient yields, particularly due to competition between product formation and cellular carbon demands across different temporal phases of growth. This study investigates how circadian clock regulation impacts carbon partitioning between storage, growth, and product synthesis in Synechococcus elongatus PCC 7942, and provides insights that suggest potential strategies for enhanced bioproduction. Results After entrainment to light-dark cycles, PCC 7942 cultures transitioned to constant light revealed distinct temporal patterns in sucrose production, exhibiting three-fold higher productivity during subjective night compared to subjective day despite moderate down-regulation of genes from the photosynthetic apparatus. This enhanced productivity coincided with reduced glycogen accumulation and halted cell division at subjective night time, suggesting temporal separation of competing processes. Transcriptome analysis revealed coordinated circadian clock-driven adjustment of the cell cycle and rewiring of energy and carbon metabolism, with over 300 genes showing differential expression across four time points. The subjective night was characterized by altered expression of cell division-related genes and reduced expression of genes involved in glycogen synthesis, while showing upregulation of glycogen degradation pathways, alternative electron flow components, the pentose phosphate pathway, and oxidative decarboxylation of pyruvate. These molecular changes created favorable conditions for product formation through enhanced availability of major sucrose precursors (glucose-1-phosphate and fructose-6-phosphate) and maintained redox balance through multiple mechanisms. Conclusions Our analysis of circadian regulatory rewiring of carbon metabolism and redox balancing suggests two potential approaches that could be developed for improving cyanobacterial bioproduction: leveraging natural circadian rhythms for optimizing cultivation conditions and timing of pathway induction, and engineering strains that mimic circadian-driven metabolic shifts through controlled carbon flux redistribution and redox rebalancing. While these strategies remain to be tested, they could theoretically improve the efficiency of photosynthetic bioproduction by enabling better temporal separation between cell growth, carbon storage accumulation, and product synthesis phases.
The composition of biomass determines its suitability for different applications within a biorefinery system. The proportion of the major biomass fractions (sugar, cellulose, hemicellulose and lignin) may vary in different sugarcane genotypes and growth environments and different parts of the plant. This study investigated the composition of mature and immature internodes, roots and mature leaves of sugarcane. Internodes were found to have a significantly larger alcohol-soluble component than leaves and roots. The primary difference between the immature and mature internodes was the ratio of soluble sugars. In mature tissues, sucrose content was significantly higher, whereas in immature internodal tissues there was lower sucrose and heightened concentrations of reducing sugars. Carbon (C) partitioning in leaf tissues was characterised by low levels of soluble components and high “other” and cell wall fractions. Root tissue had low ratios of soluble fractions relative to their cell wall contents, indicating a lack of storage of soluble carbon. There was no significant difference in the ratio of the major cell wall fractions between the major organ types. Characterisation of individual non-cellulosic monomers indicated leaf and root tissues had significantly higher arabinose and galactose fractions. Significantly larger proportions of syringyl lignin compounds and the hydroxycinnamic compound, p-coumaric acid were observed in mature internodal tissues compared to the other tissue types. Tissue-specific differences in composition were shown to greatly affect the recalcitrance of the cell wall to enzymatic saccharification. Overall, this study displayed clear evidence of the differential partitioning of C throughout the sugarcane plant in specific organs. These organ-specific differences have major implications in their utility as a bioproduct feedstock. For example, the inclusion of trash (leaves) with the culms (internodes) may alter processing efficiency.
Improving seed oil yield is essential for developing Camelina sativa as a sustainable biofuel crop. Fatty acid synthesis depends on the production of acetyl-CoA from photosynthetically derived sugars. Trehalose 6-phosphate (T6P), a proxy for sucrose availability, can link sugar status to plant growth and development. Synthesised by trehalose 6-phosphate synthase (TPS) from UDP-glucose and glucose-6-phosphate, T6P plays a regulatory role in metabolism. Our previous studies on Arabidopsis transgenic lines constitutively expressing the E. coli otsA (encoding TPS) showed increased T6P levels and seed triacylglycerol, along with stunted growth. In the present study we express otsA in camelina under the control of a seed-specific Phaseolin promoter. Seeds of the resulting transgenic lines accumulated high levels of T6P, and a 15%–20% increase in total fatty acids and triacylglycerol compared to wild-type. Molecular analysis showed the transgenic seeds had reduced SnRK1 activity, elevated WRI1 protein levels, and increased the levels of WRI1 and its target genes, along with enhanced rates of fatty acid synthesis that increased seed weights relative to wild type. Notably, the increase in oil did not affect seed protein levels but did reduce the soluble metabolite fraction. Crucially, seed-specific expression of otsA mitigated the growth defects associated with constitutive otsA expression, and the transgenic lines showed normal seed development and germination. These findings demonstrate that targeted T6P modulation via seed-specific otsA expression is an effective metabolic engineering strategy to boost oil production in camelina and potentially in other oilseed crops and bioenergy crops such as energycane, sorghum and miscanthus.
Triacetic acid lactone (TAL) can be microbially produced and further chemically upgraded to several high-value chemicals. In this work, several acidic and basic ion-exchange resins and activated charcoal were evaluated for their ability to adsorb microbially produced TAL. Activated charcoal and a weak base resin, Dowex 66, showed similar TAL adsorption capacity of 0.18 ± 0.002 g/g. At 15% w/v activated charcoal, about 98% of TAL present in fermentation broth could be adsorbed. Further, ethanol washing allowed recovery of 72% of adsorbed TAL. A biorefinery producing TAL from sucrose was designed, simulated, and evaluated (through technoeconomic analysis) under uncertainty, for an estimated TAL minimum product selling price (MPSP) of $4.27/kg [$3.71−4.94/kg; 5th-95th percentiles] for the current state of technology and $2.83/kg [$2.46–3.29/kg] following potential near-term improvements to fermentation. Thus, this work provides an adsorptive process to recover microbially produced TAL that can be chemically upgraded to several industrial products.
Bioenergy sorghum is a low-input, drought-resilient, deep-rooting annual crop that has high biomass yield potential enabling the sustainable production of biofuels, biopower, and bioproducts. Bioenergy sorghum’s 4-5 m stems account for ~80% of the harvested biomass. Stems accumulate high levels of sucrose that could be used to synthesize bioethanol and useful biopolymers if information about stem cell-type gene expression and regulation was available to enable engineering. To obtain this information, Laser Capture Microdissection (LCM) was used to isolate and collect transcriptome profiles from five major cell types that are present in stems of the sweet sorghum Wray. Transcriptome analysis identified genes with cell-type specific and cell-preferred expression patterns that reflect the distinct metabolic, transport, and regulatory functions of each cell type. Analysis of cell-type specific gene regulatory networks (GRNs) revealed that unique TF families contribute to distinct regulatory landscapes, where regulation is organized through various modes and identifiable network motifs. Cell-specific transcriptome data was combined with a stem developmental transcriptome dataset to identify the GRN that differentially activates the secondary cell wall (SCW) formation in stem xylem sclerenchyma and epidermal cells. The cell-type transcriptomic dataset provides a valuable source of information about the function of sorghum stem cell types and GRNs that will enable the engineering of bioenergy sorghum stems.
Non-typhoidal Salmonella (NTS) is a major global health concern that often causes bloodstream infections in areas of the world affected by malnutrition and comorbidities such as HIV and malaria. Developing a strategy to control the emergence and spread of highly invasive and antimicrobial resistant NTS isolates requires a comprehensive analysis of epidemiological factors and molecular pathogenesis. Here, we characterize 11 NTS isolates that caused bloodstream infections in pediatric patients in Siaya, Kenya from 2003–2010. Nine isolates were identified as S . Typhimurium sequence type 313 while the other two were S . Enteritidis. Comprehensive genotypic and phenotypic analyses were performed to compare these isolates to those previously identified in sub-Saharan Africa. We identified a S . Typhimurium isolate referred to as UGA14 that displayed novel plasmid, pseudogene and resistance features as compared to other isolates reported from Africa. Notably, UGA14 is able to ferment both lactose and sucrose due to the acquisition of insertion elements on the pKST313 plasmid. These findings show for the first time the co-evolution of plasmid-mediated lactose and sucrose metabolism along with cephalosporin resistance in NTS further elucidating the evolutionary mechanisms of invasive NTS phenotypes. These results further support the use of combined genomic and phenotypic approaches to detect and characterize atypical NTS isolates in order to advance biosurveillance efforts that inform countermeasures aimed at controlling invasive and antimicrobial resistant NTS.
Introduction Brain development involves several critical stages, such as proliferation, neuronal migration, axonal pathfinding, and connection formation. Sevoflurane, a γ -aminobutyric acid (GABA) receptor agonist, is widely used as an inhaled general anesthetic. However, its impact on brain development has raised increasing concerns, particularly regarding prenatal exposure. This study aims to investigate the effects of prenatal sevoflurane exposure (PSE) at different cortical stages, focusing on its impact on the migration of glutamatergic and GABAergic neurons and neuronal behavior in offspring. Methods PSE was administered at two critical prenatal stages: embryonic day (E) 12.5 and E18.5. Double in situ hybridization was used to identify the coexpression of GABA receptors in Pax6- and Mash1-positive cells in the forebrain. The radial migration of glutamatergic neurons and the tangential migration of GABAergic neurons were analyzed. Behavioral tests, including the open-field test, elevated plus-maze test, forced swim test, tail suspension test, sucrose preference test, and Morris water maze, were performed on offspring to assess anxiety-like behaviors, depression, and learning and memory impairments. Results PSE inhibits the radial migration of glutamatergic neurons and promotes the tangential migration of GABAergic neurons. Specifically, early exposure (E12.5) inhibited the expression of the Pax6–Tbr2–Tbr1 cascade and the radial migration of Tbr1 in the ventral prefrontal cortex (PFC), whereas late exposure (E18.5) inhibited this process on the dorsal side. In addition, offspring mice with PSE exhibited increased anxiety-like behaviors, rather than depression, as demonstrated by reduced time spent in the center of the open-field test and in the open arms of the elevated plus-maze test. No significant differences were observed in the forced swim test, tail suspension test, or sucrose preference test. Furthermore, learning and memory impairments were observed in the Morris water maze. Conclusion Our results indicate that PSE at E12.5 and E18.5 leads to abnormalities in the migration of glutamatergic and GABAergic neurons, affecting long-term anxiety-like behaviors and causing learning and memory impairments in offspring mice.
Photosynthates such as glucose, sucrose, and some of their derivatives play dual roles as metabolic intermediates and signaling molecules that influence plant cell metabolism. Such sugars provide substrates for de novo fatty acid (FA) biosynthesis. However, compared with the well-defined examples of sugar signaling in starch and anthocyanin synthesis, until recently relatively little was known about the role of signaling in regulating FA and lipid biosynthesis. Recent research progress shows that trehalose 6-phosphate and 2-oxoglutarate (2-OG) play direct signaling roles in the regulation of FA biosynthesis by modulating transcription factor stability and enzymatic activities involved in FA biosynthesis. Specifically, mechanistic links between sucrose non-fermenting−1–related protein kinase 1 (SnRK1)–mediated trehalose 6-phosphate (T6P) sensing and its regulation by phosphorylation of WRI1 stability, diacylglycerol acyltransferase 1 (DGAT1) enzyme activity, and of 2-OG–mediated relief of inhibition of acetyl-CoA carboxylase (ACCase) activity by protein PII are exemplified in detail in this review.
Seed size is a key trait affecting evolution and agronomic performance by influencing seedling establishment in natural populations and crop yields. The Arabidopsis thaliana Seed Size QTL1 (SSQ1) locus explains 10–15% of the variation in seed size. We report here that the causal gene for this locus is Tetratricopeptide Repeat Protein 2 (TPR2), which encodes a co-chaperone. Expressing TPR2 across ecotypes and genotypes showed consistent dosage effects. Each additional TPR2Col-0 allele increased seed mass and volume by 10–14% with high reliability in Col-0, Sha, Tsu-1, and tsu2 genetic backgrounds. Reciprocal genetic crosses indicated that this locus acts maternally, consistent with female sporophytic or female gametophytic mutations. To elucidate how TPR2 regulates seed size, the biomass composition of seeds was measured. While oil content remained unchanged, sucrose levels were markedly elevated in TPR2Col-0 transformant lines and reduced in tpr2 mutants. Interestingly, heterologous expression of TPR2Col-0 across genetic backgrounds increased seed protein accumulation by 18% on average. Based on these changes in sucrose and protein levels, potential modes of action for TPR2 are discussed.
In a series of experiments using sugar beets, researchers investigated the effects of varying cycles lengths on growth (0.37 hr to 48 hr). Each cycle was equally divided into a light and dark period so that each treatment regardless of cycle length received the same amount of light over the 17 weeks of the experiment. Two growth parameters were used to evaluate the effects of cycle length, total fresh weight and sucrose content of the storage root. Both parameters showed very similar responses in that under long cycles (12 hr or greater) growth was normal, whereas plants growing under shorter cycle periods were progressively inhibited. Minimum growth occurred at a cycle period of 0.75 hr. The yield at the 0.75 hr cycle, where was at a minimum, for total fresh weight was only 51 percent compared to the 24 hr cycle. The yield of sucrose was even more reduced at 41 percent of the 24 hr cycle.
This study investigated the effects of bilateral injections of a serotonin (5-HT) receptor agonist into the lateral parabrachial nucleus on the intake of NaCl and water induced by 24-h water deprivation or by sodium depletion followed by 24 h of sodium deprivation (injection of the diuretic furosemide plus 24 h of d sodium-deficient diet). Rats had stainless steel cannulas implanted bilaterally into the LPBN. Bilateral LPBN injections of the serotonergic 5-HT(1/2) receptor antagonist methysergide (4 micro-g/200 nl at each site) increased hypertonic NaCl intake when tested 24 h after sodium depletion and after 24 h of water deprivation. Water intake also increased after bilateral injections of methysergide into the LPBN. In contrast, the intake of a palatable solution (0.06 M sucrose) under body fluid-replete conditions was not changed after bilateral LPBN methysergide injections. The results show that serotonergic mechanisms in the LPBN modulate water and sodium intake induced by volume depletion and sodium loss. The finding that sucrose intake was not affected by LPBN serotonergic blockade suggests that the effects of the methysergide treatment on the intakes of water and NaCl are not due to a mechanism producing a nonspecific enhancement of all ingestive behaviors.
Ethylenediaminetetraacetic acid (EDTA), at concentrations higher than 1.0 millimolar, is phytotoxic to etiolated seedlings of Pisum sativum. Substantial vascular exudation from pea epicotyls could be obtained without tissue damage at 0.5 millimolar EDTA if the solution was buffered at pH 7.5 with sodium N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid. Treated seedlings exuded 950 micrograms (leucine equivalents) of ninhydrin-positive material per day and 870 micrograms (glucose equivalents) of anthrone-positive material per day. Amino acid analysis showed the exudate to have glutamine as the major amido nitrogen containing compound and sucrose was shown to be the major sugar. Radiolabeled tryptophan and sucrose applied to cotyledons were transferred through the epicotyl and into the collection medium. The pH profile for exudation shows half maximal exudation at pH 7.2, indicating the promotion of exudation by EDTA is probably not due simply to Ca2+ chelation.
To determine if starch statoliths do, in fact, act as gravisensors in cereal grass shoots, starch was removed from the starch statoliths by placing 45-day-old intact barley plants (Hordeum vulgare cv 'Larker') in the dark at 25 degrees C for 5 days. Evidence from staining with I2-KI, scanning electron microscopy, and transmission electron microscopy indicated that starch grains were no longer present in plastids in the pulvini of plants placed in the dark for 5 days. Furthermore, gravitropic curvature response in these pulvini was reduced to zero, even though pulvini from vertically oriented plants were still capable of elongating in response to applied auxin plus gibberellic acid. However, when 0.1 molar sucrose was fed to the dark pretreated, starch statolith-free pulvini during gravistimulation in the dark, they not only reformed starch grains in the starch-depleted plastids in the pulvini, but they also showed an upward bending response. Starch grain reformation appeared to precede reappearance of the graviresponse in these sucrose-fed pulvini. These results strongly support the view that starch statoliths do indeed serve as the gravisensors in cereal grass shoots.
Starch in pulvinus amyloplasts of barley (Hordeum vulgare cv Larker) disappears when 45-day-old, light-grown plants are given 5 days of continuous darkness. The effect of this loss on the pulvinus graviresponse was evaluated by following changes in the kinetics of response during the 5-day dark period. Over 5 days of dark pretreatment, the lag to initial graviresponse and the subsequent half-time to maximum steady state bending rate increased significantly while the maximum bending rate did not change. The change in response to applied indoleacetic acid (100 micromolar) plus gibberellic acid (10 micromolar) without gravistimulation, under identical dark pretreatments, was used as a model system for the response component of gravitropism. Dark pretreatment did not change the lag to initial response following hormone application to vertical pulvini, but both the maximum bending rate and the half-time to the maximum rate were significantly reduced. Also, after dark pretreatment, significant bending responses following hormone application were observed in vertical segments with or without added sucrose, while gravistimulation produced a response only if segments were given sucrose. These results indicate that starch-filled amyloplasts are required for the graviresponse of barley pulvini and suggest that they function in the stimulus perception and signal transduction components of gravitropism.
New yeasts from the Ross Desert (dry valley area) of Antarctica include Cryptococcus socialis sp. nov. and Cryptococcus consortionis sp. nov. Cryptococcus socialis MYSW A801-3aY1 (= ATCC 56685) requires no vitamins, assimilates L-arabinose, cellobiose, D-glucuronate, maltose, melezitose, raffinose, soluble starch, sucrose, and trehalose, and may be distinguished from all other basidioblastomycetes by the combination of amylose production, cellobiose assimilation, and failure to utilize nitrate, D-galactose, myo-inositol, and mannitol. Its guanine-plus-cytosine content is 56 mol%. Cryptococcus consortionis MYSW A801-3aY92 (= ATCC 56686) requires thiamine, assimilates L-arabinose, D-glucuronate, 2-ketogluconate, salicin, succinate, sucrose, trehalose, and D-xylose, and may be distinguished from all other basidioblastomycetes by the combination of amylose production and failure to utilize nitrate, cellobiose, D-galactose, myo-inositol, and mannitol. Its guanine-plus-cytosine content is 56 mol%.