FTY720 requires vitamin B12-TCN2-CD320 signaling in astrocytes to reduce disease in an animal model of multiple sclerosis.

Vitamin B12 (B12) deficiency causes neurological manifestations resembling multiple sclerosis (MS); however, a molecular explanation for the similarity is unknown. FTY720 (fingolimod) is a sphingosine 1-phosphate (S1P) receptor modulator and sphingosine analog approved for MS therapy that can functionally antagonize S1P1. Here, we report that FTY720 suppresses neuroinflammation by functionally and physically regulating the B12 pathways. Genetic and pharmacological S1P1 inhibition upregulates a transcobalamin 2 (TCN2)-B12 receptor, CD320, in immediate-early astrocytes (ieAstrocytes; a c-Fos-activated astrocyte subset that tracks with experimental autoimmune encephalomyelitis [EAE] severity). CD320 is also reduced in MS plaques. Deficiency of CD320 or dietary B12 restriction worsens EAE and eliminates FTY720's efficacy while concomitantly downregulating type I interferon signaling. TCN2 functions as a chaperone for FTY720 and sphingosine, whose complex induces astrocytic CD320 internalization, suggesting a delivery mechanism of FTY720/sphingosine via the TCN2-CD320 pathway. Taken together, the B12-TCN2-CD320 pathway is essential for the mechanism of action of FTY720.

Assessing the efficacy and mechanisms of glycol-contaminated water treatment through floating treatment wetlands.

The growing concerns surrounding water pollution and the degradation of ecosystems worldwide have led to an increased use of nature-based solutions (NbSs). This study assessed the feasibility of using floating treatment wetlands (FTWs) as an NbS to treat propylene glycol-contaminated water and quantitatively investigated different removal pathways. With an environmentally relevant concentration of propylene glycol (1,250 mg/L), FTWs containing Acorus calamus and mixed species demonstrated the highest average glycol mass removal efficacy (99%), followed by Carex acutiformis (98%), Juncus effusus (93%), and the control group without plants (10%) after 1 week. Additional mesocosm-scale experiments with varying FTW configurations, including surface coverage to reduce evaporation and photodegradation processes, and the addition of antibiotics to inhibit microbial activity, were conducted to quantify glycol removal pathways. Mass balance analysis results revealed that microbial biodegradation (33.3-39.7%) and plant uptake (37.9-45.2%) were the primary pathways for glycol removal. Only 15.5-19.5% of the glycol removal via evaporation and photodegradation was accounted in this study, which may be attributed to the mesocosm experimental setup (static water and no wind). Aligned with the broader discussion regarding biodiversity improvements and carbon storage capacity, this study demonstrated that FTWs are an environmentally friendly and effective NbS for addressing glycol-contaminated water.

Effects of propylene glycol used at different doses in Akkaraman lambs rations on metabolism-related parameters and liver gene and protein expression during different feeding periods.

This study aimed to investigate the metabolic effects of propylene glycol (PG) over 60, 90, and 120 days in lambs. Seventy-two weaned male lambs were allocated into three groups: control (Con), PG1.5 (1.5 mL/kg live weight0.75 ), and PG3 (3 mL/kg live weight0.75 ). Blood samples were collected at the beginning and slaughter days. Biochemical parameters (glucose, triglycerides, ALT, AST, LDH, BUN, and insulin) and gene and protein levels of peroxisome proliferator activated receptor gamma (PPARγ), diacylglycerol o-acyltransferase 1 (DGAT1), carbohydrate responsive element binding protein (ChREBP), and sterol regulatory element binding transcription factor 1c (SREBP-1c) in the liver were determined. Glucose in PG1.5 was increased on Day 60, while significant differences were observed in biochemical parameters except for insulin on the 60, 90, and 120 days. Biochemical parameters such as ALT, AST, LDH, and BUN increased over time, while triglycerides decreased. DGAT1 gene and protein levels were lower, while SREBP-1c and PPARγ were higher in PG groups on Day 60. While SREBP-1c was lower in PG1.5, ChREBP was higher in PG3 on Day 90. PPARγ, DGAT1, and ChREBP were upregulated in PG3 on Day 120. Positive correlations were found between proteins. The long-term use of PG in lambs did not have detrimental effects on metabolism. The study provides valuable insights into the molecular mechanisms underlying the metabolic effects of PG in lambs, shedding light on its potential applications in lamb production.

A genome-scale metabolic model of the effect of dissolved oxygen on 1,3-propanediol fermentation by Klebsiella pneumoniae.

Although 1,3-propanediol (1,3-PD) is usually considered an anaerobic fermentation product from glycerol by Klebsiella pneumoniae, microaerobic conditions proved to be more conducive to 1,3-PD production. In this study, a genome-scale metabolic model (GSMM) specific to K. pneumoniae KG2, a high 1.3-PD producer, was constructed. The iZY1242 model contains 2090 reactions, 1242 genes and 1433 metabolites. The model was not only able to accurately characterise cell growth, but also accurately simulate the fed-batch 1,3-PD fermentation process. Flux balance analyses by iZY1242 was performed to dissect the mechanism of stimulated 1,3-PD production under microaerobic conditions, and the maximum yield of 1,3-PD on glycerol was 0.83 mol/mol under optimal microaerobic conditions. Combined with experimental data, the iZY1242 model is a useful tool for establishing the best conditions for microaeration fermentation to produce 1,3-PD from glycerol in K. pneumoniae.

Biosynthesis of 1,3-Propanediol via a New Pathway from Glucose in Escherichia coli.

1,3-Propanediol (1,3-PDO), an important dihydric alcohol, is widely used in textiles, resins, and pharmaceuticals. More importantly, it can be used as a monomer in the synthesis of polytrimethylene terephthalate (PTT). In this study, a new biosynthetic pathway is proposed to produce 1,3-PDO using glucose as a substrate and l-aspartate as a precursor without the addition of expensive vitamin B12. We introduced a 3-HP synthesis module derived from l-aspartate and a 1,3-PDO synthesis module to achieve the de novo biosynthesis. The following strategies were then pursued that included screening key enzymes, optimizing the transcription and translation levels, enhancing the precursor supply of l-aspartate and oxaloacetate, weakening the tricarboxylic acid (TCA) cycle, and blocking competitive pathways. We also used transcriptomic methods to analyze the different gene expression levels. Finally, an engineered Escherichia coli strain produced 6.41 g/L 1,3-PDO with a yield of 0.51 mol/mol glucose in a shake flask and 11.21 g/L in fed-batch fermentation. This study provides a new pathway for production of 1,3-PDO.

Predicting human neurotoxicity of propylene glycol methyl ether (PGME) by implementing in vitro neurotoxicity results into toxicokinetic modelling.

Although organic solvents have been associated with CNS toxicity, neurotoxicity testing is rarely a regulatory requirement. We propose a strategy to assess the potential neurotoxicity of organic solvents and predict solvent air concentrations that will not likely produce neurotoxicity in exposed individuals. The strategy integrated an in vitro neurotoxicity, an in vitro blood-brain barrier (BBB), and an in silico toxicokinetic (TK) model. We illustrated the concept with propylene glycol methyl ether (PGME), widely used in industrial and consumer products. The positive control was ethylene glycol methyl ether (EGME) and negative control propylene glycol butyl ether (PGBE), a supposedly non-neurotoxic glycol ether. PGME, PGBE, and EGME had high passive permeation across the BBB (permeability coefficients (Pe) 11.0 × 10-3, 9.0 × 10-3, and 6.0 × 10-3 cm/min, respectively). PGBE was the most potent in in vitro repeated neurotoxicity assays. EGME's main metabolite, methoxyacetic acid (MAA) may be responsible for the neurotoxic effects reported in humans. No-observed adverse effect concentrations (NOAECs) for the neuronal biomarker were for PGME, PGBE, and EGME 10.2, 0.07, and 79.2 mM, respectively. All tested substances elicited a concentration-dependent increase in pro-inflammatory cytokine expressions. The TK model was used for in vitro-to-in vivo extrapolation from PGME NOAEC to corresponding air concentrations (684 ppm). In conclusion, we were able to predict air concentrations that would not likely result in neurotoxicity using our strategy. We confirmed that the Swiss PGME occupational exposure limit (100 ppm) will not likely produce immediate adverse effects on brain cells. However, we cannot exclude possible long-term neurodegenerative effects because inflammation was observed in vitro. Our simple TK model can be parameterized for other glycol ethers and used in parallel with in vitro data for systematically screening for neurotoxicity. If further developed, this approach could be adapted to predict brain neurotoxicity from exposure to organic solvents.

Strategies and tools for the biotechnological valorization of glycerol to 1, 3-propanediol: Challenges, recent advancements and future outlook.

Global efforts towards decarbonization, environmental sustainability, and a growing impetus for exploiting renewable resources such as biomass have spurred the growth and usage of bio-based chemicals and fuels. In light of such developments, the biodiesel industry will likely flourish, as the transport sector is taking several initiatives to attain carbon-neutral mobility. However, this industry would inevitably generate glycerol as an abundant waste by-product. Despite being a renewable organic carbon source and assimilated by several prokaryotes, presently realizing glycerol-based biorefinery is a distant reality. Among several platform chemicals such as ethanol, lactic acid, succinic acid, 2, 3-butanediol etc., 1, 3-propanediol (1, 3-PDO) is the only chemical naturally produced by fermentation, with glycerol as a native substrate. The recent commercialization of glycerol-based 1, 3-PDO by Metabolic Explorer, France, has revived research interests in developing alternate cost-competitive, scalable and marketable bioprocesses. The current review outlines natural glycerol assimilating and 1, 3-PDO-producing microbes, their metabolic pathways, and associated genes. Later, technical barriers are carefully examined, such as the direct use of industrial glycerol as input material and genetic and metabolic issues related to microbes alleviating their industrial use. Biotechnological interventions exploited in the past five years, which can substantially circumvent these challenges, such as microbial bioprospecting, mutagenesis, metabolic, evolutionary and bioprocess engineering, including their combinations, are discussed in detail. The concluding section sheds light on some of the emerging and most promising breakthroughs which have resulted in evolving new, efficient, and robust microbial cell factories and/or bioprocesses for glycerol-based 1, 3-PDO production.

Electrochemically mediated bioconversion and integrated purification greatly enhanced co-production of 1,3-propanediol and organic acids from glycerol in an industrial bioprocess.

In this study, we show how electrochemically mediated bioconversion can greatly increase the co-production of 1,3-propanediol and organic acids from glycerol in an industrial bioprocess using a Clostridum pasteurianum mutant. Remarkably, an enhanced butyrate formation was observed due to a weakened butanol pathway of the mutant. This allowed the strain to have a higher ATP generation for an enhanced growth, higher glycerol consumption and PDO production. The PDO titer reached as high as 120.67 g/L at a cathodic current of -400 mA, which is 33% higher than that without electricity, with a concurrent increase of butyric acid by 80%. To fully recover the increased PDO and organic acids, a novel downstream process combining thin film evaporation of PDO and esterification of organic acids with ethanol was developed. This enables the efficient co-production of PDO, ethyl acetate and ethyl butyrate with a high overall carbon use of 87%.

High-level co-production of 3-hydroxypropionic acid and 1,3-propanediol from glycerol: Metabolic engineering and process optimization.

3-Hydroxypropionic acid (3-HP) and 1,3-propanediol (1,3-PDO) are value-added chemicals with versatile applications in the chemical, pharmaceutical, and food industries. Nevertheless, sustainable production of 3-HP and 1,3-PDO is often limited by the lack of efficient strains and suitable fermentation configurations. Herein, attempts have been made to improve the co-production of both metabolites through metabolic engineering of Escherichia coli and process optimization. First, the 3-HP and 1,3-PDO co-biosynthetic pathways were recruited and optimized in E. coli, followed by coupling the pathways to the transhydrogenase-mediated cofactor regeneration systems that increased cofactor availability and product synthesis. Next, pathway rebalancing and block of by-product formation significantly improved 3-HP and 1,3-PDO net titer. Subsequently, glycerol flux toward 3-HP and 1,3-PDO synthesis was maximized by removing metabolic repression and fine-tuning the glycerol oxidation pathway. Lastly, the combined fermentation process optimization and two-stage pH-controlled fed-batch fermentation co-produced 140.50 g/L 3-HP and 1,3-PDO, with 0.85 mol/mol net yield.

Insights into C1 and C3 assimilation pathways in type I methanotrophic bacterium from co-production of 1,2-propanediol and lactate.

Methanotrophic bacteria are attractive hosts for mining metabolic pathways of C1 assimilation to produce value-added products. Herein, the type I methanotroph Methylotuvimicrobium alcaliphilum 20Z was employed to explore the carbon flux from methane and methanol via the EMP pathway to produce 1,2-propanediol (1,2-PDO). The production of 1,2-PDO on methane was found to be mainly restricted by the lower carbon flux toward the EMP pathway. The co-utilization of C1 substrates and glycerol (C3) could contribute to enhance 1,2-PDO. Lactate was co-produced in much higher amounts than 1,2-PDO. This unexpected product was probably derived from lactaldehyde by inherent aldehyde dehydrogenases. The 1,2-PDO production without increased accumulation of lactate was observed via establishing the acetol-based pathway by propane utilization with the overexpression of pmoD. This is the first study to provide experimental insights into the operation of metabolic routes for 1,2-PDO and lactate co-production from C1 and C3 compounds in methanotrophs.

Mutation of rpoS is Beneficial for Suppressing Organic Acid Secretion During 1,3-Propandiol Biosynthesis in Klebsiella pneumoniae.

In this study, to reduce the formation of organic acid during 1,3-propanediol biosynthesis in Klebsiella pneumoniae, a method combining UV mutagenesis and high-throughput screening with pH color plates was employed to obtain K. pneumoniae mutants. When compared with the parent strain, the total organic acid formation by the mutant decreased, whereas 1,3-propanediol biosynthesis increased after 24 h anaerobic shake flask culture. Subsequently, genetic changes in the mutant were analyzed by whole-genome sequencing and verified by signal gene deletion. Mutation of the rpoS gene was confirmed to contribute to the regulation of organic acid synthesis in K. pneumoniae. Besides, rpoS deletion eliminated the formation of 2,3-butanediol, the main byproduct produced during 1,3-propanediol fermentation, indicating the role of rpoS in metabolic regulation in K. pneumoniae. Thus, a K. pneumoniae mutant was developed, which could produce lower organic acid during 1,3-propanediol fermentation due to an rpoS mutation in this study.

Engineering Escherichia coli for a high yield of 1,3-propanediol near the theoretical maximum through chromosomal integration and gene deletion.

Glycerol dehydratase (gdrAB-dhaB123) operon from Klebsiella pneumoniae and NADPH-dependent 1,3-propanediol oxidoreductase (yqhD) from Escherichia coli were stably integrated on the chromosomal DNA of E. coli under the control of the native-host ldhA and pflB constitutive promoters, respectively. The developed E. coli NSK015 (∆ldhA::gdrAB-dhaB123 ∆ackA::FRT ∆pflB::yqhD ∆frdABCD::cat-sacB) produced 1,3-propanediol (1,3-PDO) at the level of 36.8 g/L with a yield of 0.99 mol/mol of glycerol consumed when glucose was used as a co-substrate with glycerol. Co-substrate of glycerol and cassava starch was also utilized for 1,3-PDO production with the concentration and yield of 31.9 g/L and 0.84 mol/mol of glycerol respectively. This represents a work for efficient 1,3-PDO production in which the overexpression of heterologous genes on the E. coli host genome devoid of plasmid expression systems. Plasmids, antibiotics, IPTG, and rich nutrients were omitted during 1,3-PDO production. This may allow a further application of E. coli NSK015 for the efficient 1,3-PDO production in an economically industrial scale. KEY POINTS:  â€¢ gdrAB-dhaB123 and yqhD were overexpressed in E. coli devoid of a plasmid system • E. coli NSK015 produced a high yield of 1,3-PDO at 99% theoretical maximum • Cassava starch was alternatively used as substrate for economical 1,3-PDO production.

Enhancing the capability of Klebsiella pneumoniae to produce 1, 3-propanediol by overexpression and regulation through CRISPR-dCas9.

Klebsiella pneumoniae is a common strain of bacterial fermentation to produce 1, 3-propanediol (1, 3-PDO). In general, the production of 1, 3-PDO by wild-type K. pneumoniae is relatively low. Therefore, a new gene manipulation of K. pneumoniae was developed to improve the production of 1, 3-PDO by overexpressing in the reduction pathway and attenuating the by-products in the oxidation pathway. Firstly, dhaB and/or dhaT were overexpressed in the reduction pathway. Considering the cost of IPTG, the constitutive promoter P32 was selected to express the key gene. By comparing K.P. pET28a-P32-dhaT with the original strain, the production of 1, 3-PDO was increased by 19.7%, from 12.97 to 15.53 g l-1 (in a 250 ml shaker flask). Secondly, three lldD and budC regulatory sites were selected in the by-product pathway, respectively, using the CRISPR-dCas9 system, and the optimal regulatory sites were selected following the 1, 3-PDO production. As a result, the 1, 3-PDO production by K.P. L1-pRH2521 and K.P. B3-pRH2521 reached up to 19.16 and 18.74 g l-1 , which was increased by 47.7% and 44.5% respectively. Overexpressing dhaT and inhibiting expression of lldD and budC were combined to further enhance the ability of K. pneumoniae to produce 1, 3-PDO. The 1, 3-PDO production by K.P. L1-B3-PRH2521-P32-dhaT reached 57.85 g l-1 in a 7.5 l fermentation tank (with Na+ neutralizer), which is higher than that of the original strain. This is the first time that the 1, 3-PDO production was improved in K. pneumoniae by overexpressing the key gene and attenuating by-product synthesis in the CRISPR-dCas9 system. This study reports an efficient approach to regulate the expression of genes in K. pneumoniae to increase the 1, 3-PDO production, and such a strategy may be useful to modify other strains to produce valuable chemicals.

Polymeric Drug Delivery System Based on Pluronics for Cancer Treatment.

Pluronic polymers (pluronics) are a unique class of synthetic triblock copolymers containing hydrophobic polypropylene oxide (PPO) and hydrophilic polyethylene oxide (PEO) arranged in the PEO-PPO-PEO manner. Due to their excellent biocompatibility and amphiphilic properties, pluronics are an ideal and promising biological material, which is widely used in drug delivery, disease diagnosis, and treatment, among other applications. Through self-assembly or in combination with other materials, pluronics can form nano carriers with different morphologies, representing a kind of multifunctional pharmaceutical excipients. In recent years, the utilization of pluronic-based multi-functional drug carriers in tumor treatment has become widespread, and various responsive drug carriers are designed according to the characteristics of the tumor microenvironment, resulting in major progress in tumor therapy. This review introduces the specific role of pluronic-based polymer drug delivery systems in tumor therapy, focusing on their physical and chemical properties as well as the design aspects of pluronic polymers. Finally, using newer literature reports, this review provides insights into the future potential and challenges posed by different pluronic-based polymer drug delivery systems in tumor therapy.

Isolation and characterization of a newly identified Clostridium butyricum strain SCUT343-4 for 1,3-propanediol production.

A novel 1,3-propanediol (1,3-PDO) producing strain was isolated and identified as Clostridium butyricum with respect to its morphological and physiological characteristics, as well as 16S rDNA. The results of substrates test and stress tolerance indicated that C. butyricum SCUT343-4 could produce 1,3-PDO efficiently from glycerol. The optimal fermentation conditions were determined to be 5 g/L yeast extract at 37 °C and pH 6.5. To fully evaluate its 1,3-PDO production capacity, different cultivation strategies have been implemented. The highest 1,3-PDO concentration obtained for batch and fed-batch fermentation were 51.64 and 61.30 g/L, respectively. Immobilized cell fermentation in fibrous-bed bioreactor was also performed, and the concentration of 1,3-PDO further increased to 86 g/L with a yield of 0.52 g/g. In addition, the 1,3-PDO productivity reached 4.20 g/L h, which is the highest level reported for C. butyricum, demonstrating the potential of C. butyricum SCUT343-4 for 1,3-PDO production from glycerol.

Anaerobic Growth of Listeria monocytogenes on Rhamnose Is Stimulated by Vitamin B12 and Bacterial Microcompartment-Dependent 1,2-Propanediol Utilization.

The foodborne pathogen Listeria monocytogenes can form proteinaceous organelles called bacterial microcompartments (BMCs) that optimize the utilization of substrates, such as 1,2-propanediol, and confer an anaerobic growth advantage. Rhamnose is a deoxyhexose sugar abundant in a range of environments, including the human intestine, and can be degraded in anaerobic conditions into 1,2-propanediol, next to acetate and lactate. Rhamnose-derived 1,2-propanediol was found to link with BMCs in some human pathogens such as Salmonella enterica, but the involvement of BMCs in rhamnose metabolism and potential physiological effects on L. monocytogenes are still unknown. In this study, we first test the effect of rhamnose uptake and utilization on anaerobic growth of L. monocytogenes EGDe without or with added vitamin B12, followed by metabolic analysis. We show that vitamin B12-dependent activation of pdu stimulates metabolism and anaerobic growth of L. monocytogenes EGDe on rhamnose via 1,2-propanediol degradation into 1-propanol and propionate. Transmission electron microscopy of pdu-induced cells shows that BMCs are formed, and additional proteomics experiments confirm expression of pdu BMC shell proteins and enzymes. Finally, we discuss the physiological effects and energy efficiency of L. monocytogenes pdu BMC-driven anaerobic rhamnose metabolism and the impact on competitive fitness in environments such as the human intestine. IMPORTANCE Listeria monocytogenes is a foodborne pathogen causing severe illness and, as such, it is crucial to understand the molecular mechanisms contributing to its survival strategy and pathogenicity. Rhamnose is a deoxyhexose sugar abundant in a range of environments, including the human intestine, and can be degraded in anaerobic conditions into 1,2-propanediol. In our previous study, the utilization of 1,2-propanediol (pdu) in L. monocytogenes was proved to be metabolized in bacterial microcompartments (BMCs), which are self-assembling subcellular proteinaceous structures and analogs of eukaryotic organelles. Here, we show that the vitamin B12-dependent activation of pdu stimulates metabolism and anaerobic growth of L. monocytogenes EGDe on rhamnose via BMC-dependent 1,2-propanediol utilization. Combined with metabolic and proteomics analysis, our discussion on the physiological effects and energy efficiency of BMC-driven rhamnose metabolism shed new light to understand the impact on L. monocytogenes competitive fitness in ecosystems such as the human intestine.

Immobilized lipase-catalyzed transesterification for synthesis of biolubricant from palm oil methyl ester and trimethylolpropane.

The present study reports the effects of three commercial immobilized lipases namely Novozyme 435 from Candida antarctica lipase B (CALB), Lipozyme TL IM from Thermomyces lanuginosus and Lipozyme RM IM from Rhizomucor miehei on the production of trimethylolpropane (TMP) ester from high oleic palm methyl ester (HO-PME) and TMP. The TMP ester is a promising base oil for biolubricants that are easily biodegradable and non-toxic to humans and the environment. Enzymatic catalysts are insensitive to free fatty acid (FFA) content, hence able to mitigate the side reactions and consequently reduce product separation cost. The potential of these enzymes to produce TMP ester in a solvent-free medium was screened at various reaction time (8, 23, 30 and 48 h), operating pressure (0.1, 0.3 and 1.0 mbar) and enzyme dosage (1, 3, 5 and 10% w/w). The reaction was conducted at a constant temperature of 70 °C and a molar ratio of 3.9:1 (HO-PME: TMP). Novozyme 435 produced the highest yield of TMP ester of 95.68 ± 3.60% under the following conditions: 23 h reaction time, 0.1 mbar operating pressure and 5% w/w of enzyme dosage. The key lubrication properties of the produced TMP ester are viscosity index (208 ± 2), pour point (- 30 ± - 2 °C), cloud point (- 15 ± - 2 °C), onset thermal degradation temperature (427.8 °C), and oxidation stability, RPVOT (42 ± 4 min). The properties of the TMP ester produced from the enzymatic transesterification are comparable to other vegetable oil-based biolubricants produced by chemical transesterification.

Unravelling the Molecular Mechanisms Underlying the Protective Effect of Lactate on the High-Pressure Resistance of Listeria monocytogenes.

Formulations with lactate as an antimicrobial and high-pressure processing (HPP) as a lethal treatment are combined strategies used to control L. monocytogenes in cooked meat products. Previous studies have shown that when HPP is applied in products with lactate, the inactivation of L. monocytogenes is lower than that without lactate. The purpose of the present work was to identify the molecular mechanisms underlying the piezo-protection effect of lactate. Two L. monocytogenes strains (CTC1034 and EGDe) were independently inoculated in a cooked ham model medium without and with 2.8% potassium lactate. Samples were pressurized at 400 MPa for 10 min at 10 °C. Samples were subjected to RNA extraction, and a shotgun transcriptome sequencing was performed. The short exposure of L. monocytogenes cells to lactate through its inoculation in a cooked ham model with lactate 1h before HPP promoted a shift in the pathogen's central metabolism, favoring the metabolism of propanediol and ethanolamine together with the synthesis of the B12 cofactor. Moreover, the results suggest an activated methyl cycle that would promote modifications in membrane properties resulting in an enhanced resistance of the pathogen to HPP. This study provides insights on the mechanisms developed by L. monocytogenes in response to lactate and/or HPP and sheds light on the understanding of the piezo-protective effect of lactate.

Hyperproduction of 3-hydroxypropionate by Halomonas bluephagenesis.

3-Hydroxypropionic acid (3HP), an important three carbon (C3) chemical, is designated as one of the top platform chemicals with an urgent need for improved industrial production. Halomonas bluephagenesis shows the potential as a chassis for competitive bioproduction of various chemicals due to its ability to grow under an open, unsterile and continuous process. Here, we report the strategy for producing 3HP and its copolymer poly(3-hydroxybutyrate-co-3-hydroxypropionate) (P3HB3HP) by the development of H. bluephagenesis. The transcriptome analysis reveals its 3HP degradation and synthesis pathways involving endogenous synthetic enzymes from 1,3-propanediol. Combing the optimized expression of aldehyde dehydrogenase (AldDHb), an engineered H. bluephagenesis strain of whose 3HP degradation pathway is deleted and that overexpresses alcohol dehydrogenases (AdhP) on its genome under a balanced redox state, is constructed with an enhanced 1.3-propanediol-dependent 3HP biosynthetic pathway to produce 154 g L-1 of 3HP with a yield and productivity of 0.93 g g-1 1,3-propanediol and 2.4 g L-1 h-1, respectively. Moreover, the strain could also accumulate 60% poly(3-hydroxybutyrate-co-32-45% 3-hydroxypropionate) in the dry cell mass, demonstrating to be a suitable chassis for hyperproduction of 3HP and P3HB3HP.

An Aldolase-Based New Pathway for Bioconversion of Formaldehyde and Ethanol into 1,3-Propanediol in Escherichia coli.

Formaldehyde (HCHO) is a reactive one-carbon compound that is interesting for biosynthesis. The assimilation of HCHO depends on the catalysis of aldolase. Here, we present a novel synthetic pathway in E. coli to convert HCHO and ethanol into 1,3-propanediol (PDO) using a deoxyribose-5-phosphate aldolase (DERA). DERA condenses HCHO and acetaldehyde to form 3-hydroxypropionaldehyde, the direct precursor of PDO formation. This new pathway opens up the possibility to synthesize an appealing C3 compound from a C1 compound and a C2 compound without carbon loss in contrast to all the other known PDO synthetic pathways where typically 30-50% of the carbons are lost as CO2 and other byproducts. The pathway is successfully demonstrated by elaborating three metabolic modules. First, DERA from Thermotoga maritima was found to be efficient for the aldol condensation and PDO production module. For the module of acetaldehyde supply from ethanol, an alcohol dehydrogenase from Hansenula polymorpha was selected. For the HCHO supply module, the control of HCHO concentration and its utilization were shown to be important for achieving the assimilation of HCHO in recombinant E. coli cells. By deleting the gene frmA for endogenous conversion of HCHO to formate and controlling HCHO at a level of about 0.6 mM, the concentration and yield of PDO were increased from initially 5.67 mM (0.43 g/L) and 0.057 mol/mol to 17.35 mM (1.32 g/L) and 0.096 mol/mol in bioconversion of ethanol and HCHO with resting E. coli cells. Further engineering of DERA and the HCHO supply module is necessary to realize the potential of this promising metabolic pathway.