Fermentable Carbon Sources Research Articles

Potential and characteristics of bio-H2 production from the garden waste of ornamental plant Malus prunifolia fruit: Comparison between ethanol-type and butyrate-type fermentations

Edible fruit waste has been studied and considered to be the ideal substrates for low-cost bio-H2 production. However, the garden waste of ornamental plant fruits, which has similar components to edible fruit, is rarely studied. In this study, the potential and characteristics of fermentative bio-H2 production from the campus ornamental plant fruit of Malus prunifolia (MP) in both ethanol- and butyrate-types were investigated. Ethanoligenens harbinense B49 and Clostridium neuense G1 were employed as the inoculums for the ethanol- and butyrate-type fermentations, respectively. The results indicated that the fruit of MP is rich in fructose and sucrose, both of strains B49 and G1 could grow well with MP powder as the carbon source for fermentation. However, a higher H2 yield of 1.77 mol-H2/mol-hexose was obtained in the butyrate-type fermentation with strain G1, which was 1.4 times of that with strain B49. C. neuense G1 was more suitable for the H2 fermentation with MP as substrate, and a H2 yield of 0.55 mol-H2/mol-hexose by strain B49 was observed in the test of buffer system exchanging from ethanol-type to the butyrate-type. In summary, garden wastes such as fruits of ornamental plants should be given more attention as low-cost fermentation substrate candidates for bio-H2 production.

Conserved mechanism of Xrn1 regulation by glycolytic flux and protein aggregation

The regulation of gene expression in eukaryotes relies largely on the action of exoribonucleases, evolutionarily conserved enzymes that digest decapped messenger RNAs in the 5’-3’ direction. The activity of Xrn1, the major yeast exoribonuclease, is regulated by targeted changes in its cellular localisation in direct response to the cell’s metabolic state. When fermentable carbon sources are available, active Xrn1 is diffusely localised in the cytosol. Upon depletion of these sources, Xrn1 is sequestered at the plasma membrane-associated protein complex, the eisosome, and becomes inactive. Although this phenomenon has been described previously, the molecular mechanisms underlying these changes remain unknown. We report that the binding of Xrn1 to the plasma membrane is subject to glycolytic flux, rather than the availability of a fermentable carbon source, is independent of TORC1 activity and requires the core eisosomal proteins Pil1 and Lsp1. We identify the SH3-like domain of the Xrn1 protein as a putative interaction domain. In addition, we show that when expressed in Saccharomyces cerevisiae, the human orthologue of Xrn1 mirrors its yeast counterpart, i.e., it segregates to the eisosome under conditions of halted glycolysis. Our results not only advance our understanding of Xrn1 regulation but also indicate that this regulatory principle is conserved from yeast to humans.

Synthetic redesign of Escherichia coli W for faster metabolism of sugarcane molasses

BackgroundSugarcane molasses, rich in sucrose, glucose, and fructose, offers a promising carbon source for industrial fermentation due to its abundance and low cost. However, challenges arise from the simultaneous utilization of multiple sugars and carbon catabolite repression (CCR). Despite its nutritional content, sucrose metabolism in Escherichia coli, except for W strain, remains poorly understood, hindering its use in microbial fermentation. In this study, E. coli W was engineered to enhance sugar consumption rates and overcome CCR. This was achieved through the integration of a synthetically designed csc operon and the optimization of glucose and fructose co-utilization pathways. These advancements facilitate efficient utilization of sugarcane molasses for the production of 3-hydroxypropionic acid (3-HP), contributing to sustainable biochemical production processes.ResultsIn this study, we addressed challenges associated with sugar metabolism in E. coli W, focusing on enhancing sucrose consumption and improving glucose-fructose co-utilization. Through targeted engineering of the sucrose utilization system, we achieved accelerated sucrose consumption rates by modulating the expression of the csc operon components, cscB, cscK, cscA, and cscR. Our findings revealed that monocistronic expression of the csc genes with the deletion of cscR, led to optimal sucrose utilization without significant growth burden. Furthermore, we successfully alleviated fructose catabolite repression by modulating the binding dynamics of FruR with the fructose PTS regulon, enabling near-equivalent co-utilization of glucose and fructose. To validate the industrial applicability of our engineered strain, we pursued 3-HP production from sugarcane molasses. By integrating heterologous genes and optimizing metabolic pathways, we achieved improvements in 3-HP titers compared to previous studies. Additionally, glyceraldehyde-3-phosphate dehydrogenase (gapA) repression aids in carbon flux redistribution, enhancing molasses conversion to 3-HP.ConclusionsDespite limitations in sucrose metabolism, the redesigned E. coli W strain, adept at utilizing sugarcane molasses, is a valuable asset for industrial fermentation. Its synthetic csc operon enhances sucrose consumption, while mitigating CCR improves glucose-fructose co-utilization. These enhancements, coupled with repression of gapA, aim to efficiently convert sugarcane molasses into 3-HP, addressing limitations in sucrose and fructose metabolism for industrial applications.

Bioprocess Optimization by Taguchi Design and Response Surface Analysis for Obtaining Active Yeast Used in Vinification

This study presents the behavior of the identified yeast strain S. cerevisiae, isolated from Busuioacă de Bohotin grapes from the Pietroasa winery, during the process of fermentation to obtain dry active yeast biomass for the winemaking process. In this respect, we promoted an optimization strategy for obtaining active dry yeast biomass. The cultivation conditions for micropilot fermentation (temperature, pH, carbon source, and nitrogen source) were selected and designed according to a Taguchi design with four factors and three levels. Reproducibility testing was conducted under specific fermentation parameters: temperature (32 °C), pH (4.5), carbon source (12%), and nitrogen source (0.7%). Following the optimization process, two combinations of cultivation parameters were selected, and one of them, based on the results, was selected for further analysis. Quantitative data were obtained, showing dry yeast biomass (DCW) at 1.39 g/100 mL and protein content at 45.57%. The active yeast was then used in the winemaking process for Tămâioasă Românească and Busuioacă de Bohotin varieties at Pietroasa winery for validation. This optimization aims to facilitate easy and rapid production of fresh wine yeast tailored to the local winemaking practices of Pietroasa winery, with real application potential in other viticultural areas, aligning with the terroir concept.

Amino acids trigger MDC-dependent mitochondrial remodeling by altering mitochondrial function.

Cells utilize numerous pathways to maintain mitochondrial homeostasis, including a recently identified mechanism that adjusts the content of the outer mitochondrial membrane (OMM) through formation of OMM-derived multilamellar domains called mitochondrial-derived compartments, or MDCs. MDCs are triggered by perturbations in mitochondrial lipid and protein content, as well as increases in intracellular amino acids. Here, we sought to understand how amino acids trigger MDCs. We show that amino acid-activation of MDCs is dependent on the functional state of mitochondria. While amino acid excess triggers MDC formation when cells are grown on fermentable carbon sources, stimulating mitochondrial biogenesis blocks MDC formation. Moreover, amino acid elevation depletes TCA cycle metabolites in yeast, and preventing consumption of TCA cycle intermediates for amino acid catabolism suppresses MDC formation. Finally, we show that directly impairing the TCA cycle is sufficient to trigger MDC formation in the absence of amino acid stress. These results demonstrate that amino acids stimulate MDC formation by perturbing mitochondrial metabolism.

Development of oil-producing microbiota and its application in fermenting distiller's grains to obtain microbial oil and ruminant feed

With the depletion of global fossil energy, it is a good choice to produce biodiesel (renewable energy) from waste biomass resources. We tried to domesticate microbiota with distiller's grains as medium, and obtained genetically stable and acid-resistant microbiota JQY1, which could convert lignocellulose into microbial oil. Amplicon sequencing results showed that Proteobacteria (94.05 %) and ascomycetes (100 %) were the main phyla, and Rhizobiaceae_unclassified (41.19 %) and Pseudomonas (24.47 %) were the dominant genera, The fungi are Wickerhamomyces and Meyerozyma, and their abundance accounts for 70.8 % and 26.32 % respectively. Using external carbon sources in the microbial body one-step fermentation of distiller's grains by microbiota JQY1 can yield a microbial oil content of 2.52 g/L. The fatty acid composition, including palmitic acid, stearic acid, oleic acid, and linoleic acid, conforms to the specifications for biodiesel raw materials set by the American ASTM. The contents of distiller's grains after fermentation were detected, in which the crude protein increased by 6.19 % and the crude fiber decreased by 10.18 %. Microbial cooperation can effectively degrade lignocellulose and transform it into microbial oil. In addition, the distiller's grains after fermentation have higher value as ruminant feed. Through one-step fermentation, we can achieve a win-win strategy of efficiently harvesting biodiesel and improving the nutritional components of distiller's grains to feed animals, which not only reduces the production cost of microbial oil, but also solves the final destination problem of distiller's grains and makes important contributions to promoting the circular economy of distiller's grains.

Study on Enhancement of Acid Production by Combination Fermentation of Caproic Acid-producing Bacteria and Lactic Acid-producing Bacteria

A new caproic acid bacteria 20-5 (Caproicibacterium sp.) was newly screened in the laboratory, and several high-quality lactic acid bacteria A2-3 ( Sporolactobacillus fermentans ), GCB-3 ( Pediococcus acidilactici ), KR-4 ( Lactiplantibacillus plantarum ), LY-5 ( Levilactobacillus brevis ), B2-1 ( Lactiplantibacillus plantarum ) were used for exploratory compound fermentation experiments. The compound fermentation was a caproic acid bacteria (20-5) alone plus four lactic acid bacteria ( A2-3, GCB-3, KR-4, LY-5, B2-1 ), the experimental ratio was set up four groups of compound ratio of 1 : 2,1 : 1,5 : 1,10 : 1. The optimum ratio of 20-5 to B2-1 was 10 : 1, and the total acid yield reached 26.75 g / L. The best caproic acid production capacity after compounding was the 10: 1 group of 20-5 and B2-1, and the caproic acid yield was 7.22 g / L. The average lactic acid content in the fermentation broth was 4.52 g / L after compounding. After compounding, the acid production of the 1 : 1 and 5 : 1 groups of 20-5 : A2-3 and the 10 : 1 group of 20-5 : B2-1 increased compared with that of single bacteria. In the other compound groups, the acid production capacity decreased after the compound operation, and the lactic acid production generally decreased, but the caproic acid production increased, indicating that caproic acid bacteria can use lactic acid as a carbon source for fermentation and acid production activities, so that caproic acid bacteria can produce caproic acid.

Bioproduction of yeast single cell oil with acute oral toxicity study intended for edible oil application.

Human nutrition and health rely on edible oils. Global demand for edible oils is expanding, necessitating the discovery of new natural oil sources subjected to adequate quality and safety evaluation. However, in contrast to other agricultural products, India's edible oil supply is surprisingly dependent on imports. The microbial oil is generated by fermentation of oleaginous yeast Rhodotorula mucilaginosa IIPL32 MTCC 25056 using biodiesel plant byproduct crude glycerol as a fermentable carbon source. Enriched with monounsaturated fatty acid, nutritional indices mapping based on the fatty acid composition of the yeast SCO, suggested its plausible use as an edible oil blend. In the present study, acute toxicity evaluation of the yeast SCO in C57BL/6 mice has been performed by randomly dividing the animals into 5 groups with 50, 300, 2000, and 5000mg/Kg yeast SCO dosage, respectively, and predicted the median lethal dose (LD50). Detailed blood biochemistry and kidney and liver histopathology analyses were also reported. The functions of the liver enzymes were also evaluated to check and confirm the anticipated toxicity. To determine cell viability and in vitro biocompatibility, the 3T3-L1 cell line and haemolysis tests were performed. The results suggested the plausible use of yeast SCO as an edible oil blend due to its non-toxic nature in mice models.

Effect of storage condition on the viability of sago effluents as carbon source in fermentation medium for bioethanol production

In Sarawak, Malaysia, approximately 237 tons/day of sago effluent is commonly discharged into nearby river due to the sago starch extraction process. Due to the high concentration of polymeric compounds, particularly starch, in sago wastewater, which petrifies easily, this condition severely pollutes the environment in the affected area. This study was conducted to determine the viability of using sago effluent as a carbon source and fermentation medium for bioethanol production which indirectly help to minimize the environmental impact as well as the economics of the sago industry. The sago effluent obtained from the local sago mill was analysed for starch content and pH profile while stored at room and cold (4°C) temperature facility. Enzymatic hydrolysis was conducted to convert the residual starch into glucose as carbon source for bioethanol fermentation. Fresh sago effluent can be stored for up to 5 days in cold temperature where the starch content remains constant. The highest starch concentration in sago effluent was 61.33 g/L, in which 50.57 g/L glucose was obtained through the enzymatic hydrolysis process. Hence 82.5% of the starch to glucose conversion yield is revealed. Then, the sago effluent hydrolysate which acts as a carbon source as well as a fermentation medium able to generate 23.14 g/L of bioethanol, displays a 91% theoretical yield of glucose to ethanol. In conclusion, the utilization of sago wastewater as feasible alternative to cheap and locally available and sustainable source of raw materials to produce bioethanol.

Optimizing Curdlan Synthesis: Engineering Agrobacterium tumefaciens ATCC31749 for Enhanced Production Using Dextrin as a Carbon Source

A key goal in current research on industrial curdlan production is the expansion of carbon sources for fermentation. In this study, a recombinant bacterial strain, sp-AmyAXCC, capable of fermenting and synthesizing curdlan using dextrin as a carbon source, was produced via heterologous expression of IPTG-inducible α-amylase from Xanthomonas campestris NRRL B-1459 in Agrobacterium tumefaciens ATCC31749. External expression of the enzyme was confirmed by western blotting, and the expression levels of exogenous proteins during the fermentation process were monitored. Additionally, the properties of the curdlan product were characterized using attenuated total reflectance-Fourier transform infrared spectroscopy and X-ray diffraction. The recombinant strain produced curdlan at a titer of 30.40 ± 0.14 g/L, gel strength of 703.5 ± 34.2 g/cm2, and a molecular weight of 3.58 × 106 Da, which is 33% greater than the molecular weight of native curdlan (2.69 × 106 Da). In the batch fermentation of sp-AmyAXCC with 12% dextrin as a carbon source, the titer of curdlan was 66.7 g/L with a yield of 0.56 g/g, and a productivity rate of 0.62 g/L/h at 108 h. The results of this study expand the substrate spectrum for Agrobacterium fermentation in curdlan production and provides guidance for further industrialization of curdlan production.

Virtually identical does not mean exactly identical: Discrepancy in energy metabolism between glucose and fructose fermentation influences the reproductive potential of yeast cells

The physiological efficiency of cells largely depends on the possibility of metabolic adaptations to changing conditions, especially on the availability of nutrients. Central carbon metabolism has an essential role in cellular function. In most cells is based on glucose, which is the primary energy source, provides the carbon skeleton for the biosynthesis of important cell macromolecules, and acts as a signaling molecule. The metabolic flux between pathways of carbon metabolism such as glycolysis, pentose phosphate pathway, and mitochondrial oxidative phosphorylation is dynamically adjusted by specific cellular economics responding to extracellular conditions and intracellular demands. Using Saccharomyces cerevisiae yeast cells and potentially similar fermentable carbon sources i.e. glucose and fructose we analyzed the parameters concerning the metabolic status of the cells and connected with them alteration in cell reproductive potential. Those parameters were related to the specific metabolic network: the hexose uptake - glycolysis and activity of the cAMP/PKA pathway - pentose phosphate pathway and biosynthetic capacities - the oxidative respiration and energy generation. The results showed that yeast cells growing in a fructose medium slightly increased metabolism redirection toward respiratory activity, which decreased pentose phosphate pathway activity and cellular biosynthetic capabilities. These differences between the fermentative metabolism of glucose and fructose, lead to long-term effects, manifested by changes in the maximum reproductive potential of cells.

A Multi-Streamline Approach for Upcycling PET into a Biodiesel and Asphalt Modifier.

The non-degradable nature of petroleum-based plastics and the dependence on petroleum-based products in daily life and production are dilemmas of human development today. We hereby developed a plastic waste upcycling process to address these challenges. A multi-stream fraction strategy was developed to process poly (ethylene terephthalate) (PET) plastics into soluble and insoluble fractions. The soluble fraction was used as a sole carbon source for microbial fermentation to produce biodiesel precursor lipids with an appreciable bioconversion yield. The insoluble fraction containing fractionated polymers was used as the asphalt binder modifiers. The downsized PET additive improved the high-temperature performance of the asphalt binder by 1 performance grade (PG) without decreasing the low-temperature PG. Subsequent SEM imaging unveiled alterations in the micromorphology induced by PET incorporation. Further FTIR and 1H NMR analysis highlighted the aromatic groups of PET polymers as a crucial factor influencing performance enhancement. The results demonstrated the multi-stream fraction as a promising approach for repurposing plastic waste to produce biodiesel and modify asphalt. This approach holds the potential to tackle challenges in fuel supply and enhance infrastructure resilience to global warming.

A novel and efficient strategy for the biodegradation of di(2-ethylhexyl) phthalate by Fusarium culmorum.

Di(2-ethylhexyl) phthalate (DEHP) is a plasticizer that is used worldwide and raises concerns because of its prevalence in the environment and potential toxicity. Herein, the capability of Fusarium culmorum to degrade a high concentration (3 g/L) of DEHP as the sole carbon and energy source in solid-state fermentation (SSF) was studied. Cultures grown on glucose were used as controls. The biodegradation of DEHP by F. culmorum reached 96.9% within 312 h. This fungus produced a 3-fold higher esterase activity in DEHP-supplemented cultures than in control cultures (1288.9 and 443.2 U/L, respectively). In DEHP-supplemented cultures, nine bands with esterase activity (24.6, 31.2, 34.2, 39.5, 42.8, 62.1, 74.5, 134.5, and 214.5 kDa) were observed by zymography, which were different from those in control cultures and from those previously reported for cultures grown in submerged fermentation. This is the first study to report the DEHP biodegradation pathway by a microorganism grown in SSF. The study findings uncovered a novel biodegradation strategy by which high concentrations of DEHP could be biodegraded using two alternative pathways simultaneously. F. culmorum has an outstanding capability to efficiently degrade DEHP by inducing esterase production, representing an ecologically promising alternative for the development of environmental biotechnologies, which might help mitigate the negative impacts of environmental contamination by this phthalate. KEY POINTS: •F. culmorum has potential to tolerate and remove di(2-ethylhexyl) phthalate (DEHP) •Solid-state fermentation is an efficient system for DEHP degradation by F. culmorum •High concentrations of DEHP induce high levels of esterase production by F. culmorum.

Efficient secretion of an enzyme cocktail in Escherichia coli for hemicellulose degradation

The use of host to secrete several hemicellulase is a cost-effective way for hemicellulose degradation. In this study, the xylose utilization gene xylAB of Escherichia coli BL21 was knocked out, and the xylanase (N20Xyl), β-xylosidase (Xys), and feruloyl esterase (FaeLam) were co-expressed in this strain. By measuring the content of reducing sugars generated by enzymatic hydrolysis of wheat bran in the fermentation supernatant, the order of the three enzymes was screened to obtain the optimal recombinant strain of E. coli BL21/∆xylAB/pDIII-2. Subsequently, fermentation conditions including culture medium, inducer concentration, induction timing, metal ions, and glycine concentration were optimized. Then, different concentrations of wheat bran and xylan were added to the fermentation medium for degradation. The results showed that the extracellular reducing sugars content reached the highest value of 33.70 ± 0.46 g/L when 50 g/L xylan was added. Besides, the scavenging rates of hydroxyl radical by the fermentation supernatant was 81.0 ± 1.41 %, and the total antioxidant capacity reached 2.289 ± 0.55. Furthermore, it showed the growth promotion effect on different lactic acid bacteria. These results provided a basis for constructing E. coli strain to efficiently degrade hemicellulose, and the strain obtained has great potential application to transform hemicellulose into fermentable carbon source.

Waste valorization through acetone-butanol-ethanol (ABE) fermentation

BackgroundBiobutanol, produced through acetone-butanol-ethanol (ABE) fermentation, has been proposed for use as a transportation fuel and for the development of butanol-based materials. However, the economic viability of ABE fermentation is heavily dependent on the cost of raw materials. MethodsThis study assesses various carbon and nitrogen sources for ABE fermentation. Carbon sources examined include local molasses (M1), Thai molasses (M2), Taiwan sugar agricultural molasses (M3), and Taiwan sugar edible molasses (M4). Nitrogen sources considered are corn steep liquor (CSL), acid-hydrolyzed black soldier fly larval meal (IA), and enzyme-hydrolyzed black soldier fly larval meal (IE). Significant findingsOur findings reveal that M2CSL, a combination of Thai molasses and corn steep liquor, serves as a cost-effective and efficient medium for ABE fermentation, resulting in a butanol concentration of 7.4 ± 3.5 g/L. Furthermore, our research identifies variations in molasses and their impact on sucrose consumption inhibition. Notably, this study underscores the potential of insect peptone derived from enzyme-hydrolyzed black soldier fly larval meal (IE) as an alternative nitrogen source, while cautioning against the use of insect peptone obtained from acid-hydrolyzed black soldier fly larval meal (IA) due to its inhibitory properties.

Utilization of glycoprotein-derived N-acetylglucosamine-L-asparagine during Enterococcus faecalis infection depends on catabolic and transport enzymes of the glycosylasparaginase locus

Enterococcus faecalis is a Gram-positive clinical pathogen causing severe infections. Its survival during infection depends on its ability to utilize host-derived metabolites, such as protein-deglycosylation products. We have identified in E. faecalis OG1RF a locus (ega) involved in the catabolism of the glycoamino acid N-acetylglucosamine-L-asparagine. This locus is separated into two transcription units, genes egaRP and egaGBCD1D2, respectively. RT-qPCR experiments revealed that the expression of the ega locus is regulated by the transcriptional repressor EgaR. Electromobility shift assays evidenced that N-acetylglucosamine-L-asparagine interacts directly with the EgaR protein, which leads to the transcription of the ega genes. Growth studies with egaG, egaB and egaC mutants confirmed that the encoded proteins are necessary for N-acetylglucosamine-L-asparagine catabolism. This glycoamino acid is transported and phosphorylated by a specific phosphotransferase system EIIABC components (OG1RF_10751, EgaB, EgaC) and subsequently hydrolyzed by the glycosylasparaginase EgaG, which generates aspartate and 6-P-N-acetyl-β-d-glucosaminylamine. The latter can be used as a fermentable carbon source by E. faecalis. Moreover, Galleria mellonella larvae had a significantly higher survival rate when infected with ega mutants compared to the wild-type strain, suggesting that the loss of N-acetylglucosamine-L-asparagine utilization affects enterococcal virulence.

Dextranase Production Using Marine Microbacterium sp. XD05 and Its Application.

Dextranase, also known as glucanase, is a hydrolase enzyme that cleaves α-1,6 glycosidic bonds. In this study, a dextranase-producing strain was isolated from water samples of the Qingdao Sea and identified as Microbacterium sp. This strain was further evaluated for growth conditions, enzyme-producing conditions, enzymatic properties, and hydrolysates. Yeast extract and sodium chloride were found to be the most suitable carbon and nitrogen sources for strain growth, while sucrose and ammonium sodium were found to be suitable carbon and nitrogen sources for fermentation. The optimal pH was 7.5, with a culture temperature of 40 °C and a culture time of 48 h. Dextranase produced by strain XD05 showed good thermal stability at 40 °C by retaining more than 70% relative enzyme activity. The pH stability of the enzyme was better under a weak alkaline condition (pH 6.0-8.0). The addition of NH4+ increased dextranase activity, while Co2+ and Mn2+ had slight inhibitory effects on dextranase activity. In addition, high-performance liquid chromatography showed that dextran is mainly hydrolyzed to maltoheptanose, maltohexanose, maltopentose, and maltootriose. Moreover, it can form corn porous starch. Dextranase can be used in various fields, such as food, medicine, chemical industry, cosmetics, and agriculture.

Molecular insights into RmcA-mediated c-di-GMP consumption: Linking redox potential to biofilm morphogenesis in Pseudomonas aeruginosa

The ability of many bacteria to form biofilms contributes to their resilience and makes infections more difficult to treat. Biofilm growth leads to the formation of internal oxygen gradients, creating hypoxic subzones where cellular reducing power accumulates, and metabolic activities can be limited. The pathogen Pseudomonas aeruginosa counteracts the redox imbalance in the hypoxic biofilm subzones by producing redox-active electron shuttles (phenazines) and by secreting extracellular matrix, leading to an increased surface area-to-volume ratio, which favors gas exchange. Matrix production is regulated by the second messenger bis-(3′,5′)-cyclic-dimeric-guanosine monophosphate (c-di-GMP) in response to different environmental cues. RmcA (Redox modulator of c-di-GMP) from P. aeruginosa is a multidomain phosphodiesterase (PDE) that modulates c-di-GMP levels in response to phenazine availability. RmcA can also sense the fermentable carbon source arginine via a periplasmic domain, which is linked via a transmembrane domain to four cytoplasmic Per-Arnt-Sim (PAS) domains followed by a diguanylate cyclase (DGC) and a PDE domain. The biochemical characterization of the cytoplasmic portion of RmcA reported in this work shows that the PAS domain adjacent to the catalytic domain tunes RmcA PDE activity in a redox-dependent manner, by differentially controlling protein conformation in response to FAD or FADH2. This redox-dependent mechanism likely links the redox state of phenazines (via FAD/FADH2 ratio) to matrix production as indicated by a hyperwrinkling phenotype in a macrocolony biofilm assay. This study provides insights into the role of RmcA in transducing cellular redox information into a structural response of the biofilm at the population level. Conditions of resource (i.e. oxygen and nutrient) limitation arise during chronic infection, affecting the cellular redox state and promoting antibiotic tolerance. An understanding of the molecular linkages between condition sensing and biofilm structure is therefore of crucial importance from both biological and engineering standpoints.

Development of a Strategy for L-Lactic Acid Production by Rhizopus oryzae Using Zizania latifolia Waste and Cane Molasses as Carbon Sources.

As a biodegradable and renewable material, polylactic acid is considered a major environmentally friendly alternative to petrochemical plastics. Microbial fermentation is the traditional method for lactic acid production, but it is still too expensive to compete with the petrochemical industry. Agro-industrial wastes are generated from the food and agricultural industries and agricultural practices. The utilization of agro-industrial wastes is an important way to reduce costs, save energy and achieve sustainable development. The present study aimed to develop a method for the valorization of Zizania latifolia waste and cane molasses as carbon sources for L-lactic acid fermentation using Rhizopus oryzae LA-UN-1. The results showed that xylose derived from the acid hydrolysis of Z. latifolia waste was beneficial for cell growth, while glucose from the acid hydrolysis of Z. latifolia waste and mixed sugars (glucose and fructose) from the acid hydrolysis of cane molasses were suitable for the accumulation of lactic acid. Thus, a three-stage carbon source utilization strategy was developed, which markedly improved lactic acid production and productivity, respectively reaching 129.47 g/L and 1.51 g/L·h after 86 h of fermentation. This work demonstrates that inexpensive Z. latifolia waste and cane molasses can be suitable carbon sources for lactic acid production, offering an efficient utilization strategy for agro-industrial wastes.

Construction of Recombinant Escherichia coli with a High L-Phenylalanine Production Yield from Glucose

L-phenylalanine is an important aromatic amino acid that is widely used in the area of feed, food additives, and pharmaceuticals. Among the different strategies of L-phenylalanine synthesis, direct microbial fermentation from raw substrates has attracted more and more attention due to its environment friendly process and low-cost raw materials. In this study, a rational designed recombinant Escherichia coli was constructed for L-phenylalanine production. Based on wild type E. coli MG1655, multilevel engineering strategies were carried out, such as directing more carbon flux into the L-phenylalanine synthetic pathway, increasing intracellular level of precursors, blocking by-product synthesis pathways and facilitating the secretion of L-phenylalanine. During 5 L fed batch fermentation, recombinant E. coli MPH-3 could produce 19.24 g/L of L-phenylalanine with a yield of 0.279 g/g glucose. To the best of our knowledge, this is one of the highest yields of L-phenylalanine producing E. coli using glucose as the sole carbon source in fed-batch fermentation.