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Cryptosporidium parvum is a waterborne and foodborne zoonotic protozoan parasite, a causative agent of moderate to severe diarrheal diseases in humans and animals. However, fully effective treatments are unavailable for medical and veterinary uses. There is a need to explore new drug targets for potential development of new therapeutics. Because C. parvum relies on anaerobic metabolism to produce ATP, fermentative enzymes in this parasite are attractive targets for exploration. In this study, we investigated the ethanol-fermentation in the parasite and characterized the basic biochemical features of a bacterial-type bifunctional aldehyde/alcohol dehydrogenase, namely CpAdhE. We also screened 3892 chemical entries from three libraries and identified 14 compounds showing >50% inhibition on the enzyme activity of CpAdhE. Intriguingly, antifungal imidazoles and unsaturated fatty acids are the two major chemical groups among the top hits. We further characterized the inhibitory kinetics of selected imidazoles and unsaturated fatty acids on CpAdhE. These compounds displayed lower micromolar activities on CpAdhE (i.e., IC50 values ranging from 0.88 to 11.02 μM for imidazoles and 8.93 to 35.33 μM for unsaturated fatty acids). Finally, we evaluated the in vitro anti-cryptosporidial efficacies and cytotoxicity of three imidazoles (i.e., tioconazole, miconazole and isoconazole). The three antifungal imidazoles exhibited lower micromolar efficacies against the growth of C. parvum in vitro (EC50 values ranging from 4.85 to 10.41 μM and selectivity indices ranging from 5.19 to 10.95). The results provide a proof-of-concept data to support that imidazoles are worth being further investigated for potential development of anti-cryptosporidial therapeutics.

In this investigation, the preparation of the chiral intermediate, (R)‐1‐(2‐chlorophenyl)‐2‐(2 H‐tetrazol‐2‐yl)ethan‐1‐ol (2 a), a precursor to the anti‐epileptic drug Cenobamate, was demonstrated using a very efficient alcohol dehydrogenase (ADH) enzyme from a simple ketone substrate. The enzyme, ADH107604 from Streptomyces coelicolor (ScADH), identified from Dr. Reddy's in‐house collection of biocatalysts, showed excellent activity against the ketone 1‐(2‐chlorophenyl)‐2‐(2 H‐tetrazol‐2‐yl)ethan‐1‐one 1, and the (R)‐alcohol product 2 a was obtained with excellent conversion and enantiomeric excess (ee) >99 %. [ABSTRACT FROM AUTHOR]

The following review article presents clinical and experimental features of alcohol-induced liver disease (ALD). Basic aspects of alcohol metabolism leading to the development of liver hepatotoxicity are discussed. ALD includes fatty liver, acute alcoholic hepatitis with or without liver failure, alcoholic steatohepatitis (ASH) leading to fibrosis and cirrhosis, and hepatocellular cancer (HCC). ALD is fully attributable to alcohol consumption. However, only 10-20% of heavy drinkers (persons consuming more than 40 g of ethanol/day) develop clinical ALD. Moreover, there is a link between behaviour and environmental factors that determine the amount of alcohol misuse and their liver disease. The range of clinical presentation varies from reversible alcoholic hepatic steatosis to cirrhosis, hepatic failure, and hepatocellular carcinoma. We aimed to (1) describe the clinico-pathology of ALD, (2) examine the role of immune responses in the development of alcoholic hepatitis (ASH), (3) propose diagnostic markers of ASH, (4) analyze the experimental models of ALD, (5) study the role of alcohol in changing the microbiota, and (6) articulate how findings in the liver and/or intestine influence the brain (and/or vice versa) on ASH; (7) identify pathways in alcohol-induced organ damage and (8) to target new innovative experimental concepts modeling the experimental approaches. The present review includes evidence recognizing the key toxic role of alcohol in ALD severity. Cytochrome p450 CYP2E1 activation may change the severity of ASH. The microbiota is a key element in immune responses, being an inducer of proinflammatory T helper 17 cells and regulatory T cells in the intestine. Alcohol consumption changes the intestinal microbiota and influences liver steatosis and liver inflammation. Knowing how to exploit the microbiome to modulate the immune system might lead to a new form of personalized medicine in ALF and ASH.

As a flooding-tolerant tree species, Taxodium distichum has been utilized in afforestation projects and proven to have important value in flooding areas. Alcohol dehydrogenase (ADH), which participates in ethanol fermentation, is essential for tolerance to the anaerobic conditions caused by flooding. In a comprehensive analysis of the ADH gene family in T. distichum, TdADHs were cloned on the basis of whole-genome sequencing, and then bioinformatic analysis, subcellular localization, and gene expression level analysis under flooding were conducted. The results show that the putative protein sequences of 15 cloned genes contained seven TdADHs and eight TdADH-like genes (one Class III ADH included) that were divided into five clades. All the sequences had an ADH_N domain, and except for TdADH-likeE2, all the other genes had an ADH_zinc_N domain. Moreover, the TdADHs in clades A, B, C, and D had a similar motif composition. Additionally, the number of TdADH amino acids ranged from 277 to 403, with an average of 370.13. Subcellular localization showed that, except for TdADH-likeD3, which was not expressed in the nucleus, the other genes were predominantly expressed in both the nucleus and cytosol. TdADH-likeC2 was significantly upregulated in all three organs (roots, stems, and leaves), and TdADHA3 was also highly upregulated under 24 h flooding treatment; the two genes might play key roles in ethanol fermentation and flooding tolerance. These findings offer a comprehensive understanding of TdADHs and could provide a foundation for the molecular breeding of T. distichum and current research on the molecular mechanisms driving flooding tolerance. [ABSTRACT FROM AUTHOR]

The objective was to analyze the influence of industrial soybean seed treatments on physiological quality throughout the storage and to evaluate the worth of the alcohol dehydrogenase (ADH) enzyme as a marker for quality monitoring. The statistical design used was completely randomized in a 2 x 5 x 5 factorial schemes involving two cultivars, five phytosanitary treatments, and five evaluation periods during storage. Before treatment, the lignin content of the seed coat of the cultivars was analyzed. The physiological quality was evaluated by the first count of germination and the germination, and the ADH enzyme was also quantified. The deterioration of soybean seeds during storage was directly linked to the type of molecule and the viscosity of the spray solution used in the treatment of seeds. The treatment of soybean seeds with predominantly aqueous solution and the use of systemic insecticidal molecules, combined with fungicides and polymers, impaired the germination of the stored seeds, and the deleterious effects became greater over time. ADH has potential as a marker of the physiological quality of treated and stored soybean seeds. [ABSTRACT FROM AUTHOR]

Cryptosporidium parvum is a waterborne and foodborne zoonotic protozoan parasite, a causative agent of moderate to severe diarrheal diseases in humans and animals. However, fully effective treatments are unavailable for medical and veterinary uses. There is a need to explore new drug targets for potential development of new therapeutics. Because C. parvum relies on anaerobic metabolism to produce ATP, fermentative enzymes in this parasite are attractive targets for exploration. In this study, we investigated the ethanol-fermentation in the parasite and characterized the basic biochemical features of a bacterial-type bifunctional aldehyde/alcohol dehydrogenase, namely CpAdhE. We also screened 3892 chemical entries from three libraries and identified 14 compounds showing >50% inhibition on the enzyme activity of CpAdhE. Intriguingly, antifungal imidazoles and unsaturated fatty acids are the two major chemical groups among the top hits. We further characterized the inhibitory kinetics of selected imidazoles and unsaturated fatty acids on CpAdhE. These compounds displayed lower micromolar activities on CpAdhE (i.e., IC 50 values ranging from 0.88 to 11.02 μM for imidazoles and 8.93 to 35.33 μM for unsaturated fatty acids). Finally, we evaluated the in vitro anti-cryptosporidial efficacies and cytotoxicity of three imidazoles (i.e., tioconazole, miconazole and isoconazole). The three antifungal imidazoles exhibited lower micromolar efficacies against the growth of C. parvum in vitro (EC 50 values ranging from 4.85 to 10.41 μM and selectivity indices ranging from 5.19 to 10.95). The results provide a proof-of-concept data to support that imidazoles are worth being further investigated for potential development of anti-cryptosporidial therapeutics. [Display omitted] [ABSTRACT FROM AUTHOR]

Background and Aims: Nebbiolo grapes are used to produce Sfursat wine, following partial dehydration. This research aimed to clarify the influence of fruit exposure to light and postharvest water loss on the concentration of green leaf volatiles (GLVs) and lipoxygenase (LOX) and alcohol dehydrogenase (ADH) activity of grapes. Methods and Results: Nebbiolo grapes from Control vines (no defoliation) (ND) and from vines defoliated at fruitset (DFS) or defoliated post‐veraison (DPV) were harvested at about 23°Brix and dehydrated at 10 and 20°C, 60% RH and air flow of 1.5 m/s. Berries were sampled at 10 and 20% mass loss (ML). Significant differences in crop yield, bunch mass and berry mass were observed. As expected, the higher the dehydration temperature, the faster the dehydration process: 20% ML at 20°C occurred between 18 and 25 days, the shortest time corresponding to ND and the longest to DFS; at 10°C, the dehydration lasted between 27 and 32 days. At 10°C, the ADH activity was almost double that at 20°C, and in DFS was much higher than in other samples. At harvest, LOX did not show any difference among the samples, while at 10°C and 10% ML, the enzyme activity increased significantly and then declined at 20% ML, especially in defoliated samples. At harvest, the total free GLVs associated with the metabolism of lipid oxidation were 9434, 7212 and 11 656 μg/kg dry weight (DW) in ND, DFS and DPV samples, respectively; the total bound GLVs lipid‐derived were 7599, 18 486 and 15 409 μg/kg DW in ND, DFS and DPV samples, respectively. During dehydration at 10°C, the ML induced ADH + LOX activity, especially in defoliated samples, but the bound GLVs, produced by defoliation, greatly decreased. Conclusions: Defoliation affected the response of Nebbiolo grapes to dehydration temperature: postharvest cold stress (10°C) and ML induced glycosylation of GLVs, alcohol formation (via ADH) and membrane oxidation (via LOX); a further stress effect was observed with leaf removal, regardless of the time of application. Significance of the Study: The timing of defoliation and postharvest dehydration temperature are significant factors to mitigate the postharvest stress response of Nebbiolo grapes. [ABSTRACT FROM AUTHOR]

As a flooding-tolerant tree species, Taxodium distichum has been utilized in afforestation projects and proven to have important value in flooding areas. Alcohol dehydrogenase (ADH), which participates in ethanol fermentation, is essential for tolerance to the anaerobic conditions caused by flooding. In a comprehensive analysis of the ADH gene family in T. distichum, TdADHs were cloned on the basis of whole-genome sequencing, and then bioinformatic analysis, subcellular localization, and gene expression level analysis under flooding were conducted. The results show that the putative protein sequences of 15 cloned genes contained seven TdADHs and eight TdADH-like genes (one Class III ADH included) that were divided into five clades. All the sequences had an ADH_N domain, and except for TdADH-likeE2, all the other genes had an ADH_zinc_N domain. Moreover, the TdADHs in clades A, B, C, and D had a similar motif composition. Additionally, the number of TdADH amino acids ranged from 277 to 403, with an average of 370.13. Subcellular localization showed that, except for TdADH-likeD3, which was not expressed in the nucleus, the other genes were predominantly expressed in both the nucleus and cytosol. TdADH-likeC2 was significantly upregulated in all three organs (roots, stems, and leaves), and TdADHA3 was also highly upregulated under 24 h flooding treatment; the two genes might play key roles in ethanol fermentation and flooding tolerance. These findings offer a comprehensive understanding of TdADHs and could provide a foundation for the molecular breeding of T. distichum and current research on the molecular mechanisms driving flooding tolerance.

Although Alzheimer's disease (AD) characterized with senile plaques and neurofibrillary tangles has been found for over 100 years, its molecular mechanisms are ambiguous. More worsely, the developed medicines targeting amyloid-beta (Aβ) and/or tau hyperphosphorylation did not approach the clinical expectations in patients with moderate or severe AD until now. This review unveils the role of a vicious cycle between Aβ-derived formaldehyde (FA) and FA-induced Aβ aggregation in the onset course of AD. Document evidence has shown that Aβ can bind with alcohol dehydrogenase (ADH) to form the complex of Aβ/ADH (ABAD) and result in the generation of reactive oxygen species (ROS) and aldehydes including malondialdehyde, hydroxynonenal and FA; in turn, ROS-derived H 2 O 2 and FA promotes Aβ self-aggregation; subsequently, this vicious cycle accelerates neuron death and AD occurrence. Especially, FA can directly induce neuron death by stimulating ROS generation and tau hyper hyperphosphorylation, and impair memory by inhibiting NMDA-receptor. Recently, some new therapeutical methods including inhibition of ABAD activity by small molecules/synthetic polypeptides, degradation of FA by phototherapy or FA scavengers, have been developed and achieved positive effects in AD transgenic models. Thus, breaking the vicious loop may be promising interventions for halting AD progression. [Display omitted] • Aβ can bind with alcohol dehydrogenase (ADH) to form the complex of Aβ/ADH. • ADH inactivation induces FA generation; in turn, FA promotes Aβ self-aggregation. • Vicious cycle between Aβ-derived FA and FA-induced Aβ assembly elicits AD onset. • Degradation of FA alleviates Aβ neurotoxicity and Alzheimer-related phenotypes. • Breaking the vicious cycle is promising interventions for halting AD progression. [ABSTRACT FROM AUTHOR]

Key message: The study shows the biochemical and enzymatic divergence between the two aldehyde-alcohol dehydrogenases of the alga Polytomella sp., shedding light on novel aspects of the enzyme evolution amid unicellular eukaryotes. Aldehyde-alcohol dehydrogenases (ADHEs) are large metalloenzymes that typically perform the two-step reduction of acetyl-CoA into ethanol. These enzymes consist of an N-terminal acetylating aldehyde dehydrogenase domain (ALDH) and a C-terminal alcohol dehydrogenase (ADH) domain. ADHEs are present in various bacterial phyla as well as in some unicellular eukaryotes. Here we focus on ADHEs in microalgae, a diverse and polyphyletic group of plastid-bearing unicellular eukaryotes. Genome survey shows the uneven distribution of the ADHE gene among free-living algae, and the presence of two distinct genes in various species. We show that the non-photosynthetic Chlorophyte alga Polytomella sp. SAG 198.80 harbors two genes for ADHE-like enzymes with divergent C-terminal ADH domains. Immunoblots indicate that both ADHEs accumulate in Polytomella cells growing aerobically on acetate or ethanol. ADHE1 of ~ 105-kDa is found in particulate fractions, whereas ADHE2 of ~ 95-kDa is mostly soluble. The study of the recombinant enzymes revealed that ADHE1 has both the ALDH and ADH activities, while ADHE2 has only the ALDH activity. Phylogeny shows that the divergence occurred close to the root of the Polytomella genus within a clade formed by the majority of the Chlorophyte ADHE sequences, next to the cyanobacterial clade. The potential diversification of function in Polytomella spp. unveiled here likely took place after the loss of photosynthesis. Overall, our study provides a glimpse at the complex evolutionary history of the ADHE in microalgae which includes (i) acquisition via different gene donors, (ii) gene duplication and (iii) independent evolution of one of the two enzymatic domains. [ABSTRACT FROM AUTHOR]

The cellulolytic bacterium Clostridium thermocellum is a promising candidate for lignocellulosic biofuel production; however, ethanol titer needs to be improved for commercialization. To understand the factors limiting ethanol titer in C. thermocellum, we developed a cell-free extract reaction (CFER) system. We demonstrated that 15 mM cellobiose could be converted, in vitro, to 25 mM ethanol and that this reaction functions both at thermophilic (55°C) and mesophilic (37°C) temperatures. Although the yield was similar to that produced by whole cells, the rate was much slower (~0.5 vs. 12 mM/h). In order to reliably quantify metabolites, rapid CFER quenching is necessary. Among the methods tested, filtration with a 3-kDa molecular weight cutoff filter proved to be the most effective. Metabolomic analysis revealed high levels of glucose-6-phosphate (G6P) and fructose-6-phosphate (F6P) in the CFER, identifying potential rate-limiting enzymes downstream of F6P. NADH was also found to accumulate in the CFER, suggesting that NADH recycling is rate-limiting. We used two complementary strategies to identify enzymes that limit metabolic flux, including feeding different substrates and supplementing with exogenous enzymes. In the enzyme addition experiment, the largest improvement was observed with the addition of yeast alcohol dehydrogenase (ADH), indicating a limitation at that reaction. The development of a CFER system for C. thermocellum, combined with detailed measurements of intermediate metabolites, allowed us to directly observe the metabolism of this organism, and suggested several potential metabolic engineering interventions for increasing ethanol titer. This demonstrates a technique that may be of general use for metabolic engineering in non-model organisms.

The main metabolic pathway of ethanol elimination occurs in the liver. Ethanol is oxidised by an alcohol dehydrogenase (ADH) to a very toxic acetaldehyde. The aldehyde is converted to acetate by an aldehyde dehydrogenase (ALDH). Studies of animals with genetic predisposition to drink ethanol indicate the relationship between spontaneous drinking of alcohol and its increased metabolism. Chronic alcohol consumption is significantly related to the risk of developing cancer in the upper gastrointestinal tract (oral cavity and oesophagus) as well as liver, pancreas, and large intestine. Studies show that the carcinogenic effect of alcohol is due to the action of acetaldehyde, which interferes in many places with DNA synthesis, influences DNA repair capacity, and binding to cellular proteins causes cellular dysfunction. The aldehyde can generate errors of gene replication. Other mechanisms involved in ethanol metabolism are associated with reactive oxygen species (ROS) resulting from induction of cytochrome P4502E1. Reactive oxygen species can directly affect DNA or by lipid oxidation resulting in breaks in the DNA chain. Both the formation and metabolism of acetaldehyde is modified by ADH and ALDH. Alcohol dehydrogenase activity is significantly higher in tumour tissues than in healthy tissues and is disproportionately higher in relation to ALDH activity. Such enzyme activity may indicate faster alcohol oxidation and significantly reduced removal of acetaldehyde in the case of cancer. As a result of the diminished acetic aldehyde reduction, its concentration may increase in the tumour tissue and thus may potentiate its carcinogenic effect.

Background: Inactive aldehyde dehydrogenase‐2 (ALDH2) is a well‐known deterrent to the development of alcohol use disorder (AUD), and however, some individuals with inactive ALDH2 do go on to develop AUD. These alcoholics are likely to have strong risk factors for the development of this disorder. Using a model of alcoholics with inactive ALDH2 (the AIA model), we investigated the unique characteristics of alcoholics with inactive ALDH2 in an attempt to identify the risk factors for AUD. In this study, we focused on comorbid psychiatric and personality disorders as potential risk factors for AUD. Methods: The subjects were 103 male alcoholics with inactive ALDH2 (AIAs), 87 age‐ and ADH1B genotype‐matched alcoholics with active ALDH2 (AAAs) and 200 age‐matched healthy men. The alcoholics were divided into 4 subgroups according to their ALDH2 and ADH1B genotypes (inactive ALDH2 vs. active ALDH2, usual ADH1B vs. superactive ADH1B). To assess the participants' comorbid psychiatric disorders, we conducted semi‐structured interviews using the Japanese translation of SSAGA version 2. We compared the prevalence of comorbid psychiatric and personality disorders among groups with different combinations of the ALDH2 and ADH1B genotypes. Results: The prevalence of attention‐deficit/hyperactivity disorder (ADHD) was significantly higher in the AIAs with usual ADH1B than in the other 3 subgroups of alcoholics. In contrast, the prevalence rates of agoraphobia and panic disorder were significantly lower in the AIAs with superactive ADH1B than in the other 3 subgroups of alcoholics. Conclusions: This study suggested that (i) ADHD is a risk factor for AUD, consistent with previous reports; (ii) agoraphobia and panic disorder may have deterrent effects against the development of AUD in individuals with inactive ALDH2, probably attributable to the similarity between the symptoms of agoraphobia and panic disorder and the adverse reactions to consumption of alcohol in subjects with inactive ALDH2. [ABSTRACT FROM AUTHOR]

Excessive alcohol consumption can cause multi-systemic diseases. Among them, alcoholic liver disease is the most frequent and serious disease. Electrolytic hydrogen water (EHW) is produced at the cathode during electrolysis of water and contains a large amount of molecular hydrogen and a low content of platinum nanoparticles with alkaline properties. In this study, we found that EHW inhibits ethanol-induced cytotoxicity by decreasing the intracellular acetaldehyde, a toxic substance produced by ethanol degradation, in hepatocyte cell lines HepG2. Analysis of the mechanism of action revealed that EHW inhibits the metabolism of ethanol to acetaldehyde by suppressing alcohol dehydrogenase. EHW also promotes the metabolism of acetaldehyde to acetic acid by activating aldehyde dehydrogenase, which plays to reduce aldehyde toxicity and intracellular reactive oxygen species in HepG2 cells. These functions were correlated with the concentration of molecular hydrogen in EHW, and were abolished by degassing treatment, suggesting that molecular hydrogen may contribute as a functional factor in the suppression of ethanol-induced hepatocellular damage. Furthermore, hydrogen water with high dissolved hydrogen molecule showed the same hepatocellular protective effect against ethanol as the EHW. These results suggest that EHW may be useful in the prevention of alcoholic liver disease.

Aim: Paclitaxel (PTX) has no clinically available oral formulations. Cetyl alcohol is metabolized by alcohol dehydrogenase and aldehyde dehydrogenase that are overexpressed in cancer cells. So, PTX-encapsulated core–shell nanoparticle of cetyl alcohol (PaxSLN) could target the cancer cells through oral route. Materials & methods: PaxSLN was synthesized using microemulsion template. Efficiency of PaxSLN was evaluated by ALDEFLUOR™, multicellular tumor spheroid formation inhibition assays and CT26 colorectal carcinoma animal model. Pharmacokinetics and biodistribution studies were done in Sprague Dawley rats. Results: PTX was encapsulated at the core of approximately 78 nm PaxSLN. PaxSLN targeted aldehyde dehydrogenase overexpressing cells. Its oral bioavailability was approximately 95% and chemotherapeutic efficacy was better than Taxol® and nab-PTX. Conclusion: A novel oral nanoformulation of PTX was developed. [ABSTRACT FROM AUTHOR]

The following review article presents clinical and experimental features of alcohol-induced liver disease (ALD). Basic aspects of alcohol metabolism leading to the development of liver hepatotoxicity are discussed. ALD includes fatty liver, acute alcoholic hepatitis with or without liver failure, alcoholic steatohepatitis (ASH) leading to fibrosis and cirrhosis, and hepatocellular cancer (HCC). ALD is fully attributable to alcohol consumption. However, only 10−20% of heavy drinkers (persons consuming more than 40 g of ethanol/day) develop clinical ALD. Moreover, there is a link between behaviour and environmental factors that determine the amount of alcohol misuse and their liver disease. The range of clinical presentation varies from reversible alcoholic hepatic steatosis to cirrhosis, hepatic failure, and hepatocellular carcinoma. We aimed to (1) describe the clinico-pathology of ALD, (2) examine the role of immune responses in the development of alcoholic hepatitis (ASH), (3) propose diagnostic markers of ASH, (4) analyze the experimental models of ALD, (5) study the role of alcohol in changing the microbiota, and (6) articulate how findings in the liver and/or intestine influence the brain (and/or vice versa) on ASH; (7) identify pathways in alcohol-induced organ damage and (8) to target new innovative experimental concepts modeling the experimental approaches. The present review includes evidence recognizing the key toxic role of alcohol in ALD severity. Cytochrome p450 CYP2E1 activation may change the severity of ASH. The microbiota is a key element in immune responses, being an inducer of proinflammatory T helper 17 cells and regulatory T cells in the intestine. Alcohol consumption changes the intestinal microbiota and influences liver steatosis and liver inflammation. Knowing how to exploit the microbiome to modulate the immune system might lead to a new form of personalized medicine in ALF and ASH.

To produce bioethanol from model cyanobacteria such as Synechocystis, a two gene cassette consisting of genes encoding pyruvate decarboxylase (PDC) and alcohol dehydrogenase (ADH) are required to transform pyruvate first to acetaldehyde and then to ethanol. However the partition of pyruvate to ethanol comes at a cost, a reduction in biomass and pyruvate availability for other metabolic processes. Hence strategies to divert flux to ethanol as a biofuel in Synechocystis are of interest. PDC from Zymobacter palmae (ZpPDC) has been reported to have a lower Km then the Zymomonas mobilis PDC (ZmPDC), which has traditionally been used in metabolic engineering constructs. The Zppdc gene was combined with the native slr1192 alcohol dehydrogenase gene (adhA) in an attempt to increase ethanol production in the photoautotrophic cyanobacterium Synechocystis sp. PCC 6803 over constructs created with the traditional Zmpdc. Native (Zppdc) and codon optimized (ZpOpdc) versions of the ZpPDC were cloned into a construct where pdc expression was controlled via the psbA2 light inducible promoter from Synechocystis sp. PCC 6803. These constructs were transformed into wildtype Synechocystis sp. PCC 6803 for expression and ethanol production. Ethanol levels were then compared with identical constructs containing the Zmpdc. While strains with the Zppdc (UL071) and ZpOpdc (UL072) constructs did produce ethanol, levels were lower compared to a control strain (UL070) expressing the pdc from Zymomonas mobilis. All constructs demonstrated lower biomass productivity illustrating that the flux from pyruvate to ethanol has a major effect on biomass and ultimately overall biofuel productivity. Thus the utilization of a PDC with a lower Km from Zymobacter palmae unusually did not result in enhanced ethanol production in Synechocystis sp. PCC 6803.

The spectrum of alcoholic liver disease (ALD) is broad and includes alcoholic fatty liver, alcoholic steatohepatitis, alcoholic hepatitis, alcoholic fibrosis, alcoholic cirrhosis, and alcoholic hepatocellular carcinoma, best explained as a five-hit sequelae of injurious steps. ALD is not primarily the result of malnutrition as assumed for many decades but due to the ingested alcohol and its metabolic consequences although malnutrition may marginally contribute to disease aggravation. Ethanol is metabolized in the liver to the heavily reactive acetaldehyde via the alcohol dehydrogenase (ADH) and the cytochrome P450 isoform 2E1 of the microsomal ethanol-oxidizing system (MEOS). The resulting disturbances modify not only the liver parenchymal cells but also non-parenchymal cells such as Kupffer cells (KCs), hepatic stellate cells (HSCs), and liver sinusoidal endothelial cells (LSECs). These are activated by acetaldehyde, reactive oxygen species (ROS), and endotoxins, which are produced from bacteria in the gut and reach the liver due to gut leakage. A variety of intrahepatic signaling pathways and innate or acquired immune reactions are under discussion contributing to the pathogenesis of ALD via the five injurious hits responsible for disease aggravation. As some of the mechanistic steps are based on studies with in vitro cell systems or animal models, respective proposals for humans may be considered as tentative. However, sufficient evidence is provided for clinical risk factors that include the amount of alcohol used daily for more than a decade, gender differences with higher susceptibility of women, genetic predisposition, and preexisting liver disease. In essence, efforts within the last years were devoted to shed more light in the pathogenesis of ALD, much has been achieved but issues remain to what extent results obtained from experimental studies can be transferred to humans.

NAD(P)-dependent group III alcohol dehydrogenases (ADHs), well known as iron-activated enzymes, generally lose their activities under aerobic conditions due to their oxygen-sensitivities. In this paper, we expressed an extremely thermostable group III ADH from the hyperthermophilic archaeon Pyrococcus horikoshii OT3 ( PhADH) heterologously in Escherichia coli. When purified from a culture medium containing nickel, the recombinant PhADH (Ni- PhADH) contained 0.85 ± 0.01 g-atoms of nickel per subunit. Ni- PhADH retained high activity under aerobic conditions (9.80 U mg), while the enzyme expressed without adding nickel contained 0.46 ± 0.01 g-atoms of iron per subunit and showed little activity (0.27 U mg). In the presence of oxygen, the activity of the Fe-reconstituted PhADH prepared from the Ni- PhADH was gradually decreased, whereas the Ni-reconstituted PhADH maintained enzymatic activity. These results indicated that PhADH with bound nickel ion was stable in oxygen. The activity of the Ni-reconstituted PhADH prepared from the expression without adding nickel was significantly lower than that from the Ni- PhADH, suggesting that binding a nickel ion to PhADH in this expression system contributed to protecting against inactivation during the expression and purification processes. Unlike other thermophilic group III ADHs, Ni- PhADH showed high affinity for NAD(H) rather than NADP(H). Furthermore, it showed an unusually high k value toward aldehyde reduction. The activity of Ni- PhADH for butanal reduction was increased to 60.7 U mg with increasing the temperature to 95 °C. These findings provide a new strategy to obtain oxygen-sensitive group III ADHs. [ABSTRACT FROM AUTHOR]

A multispectroscopic exploration was employed to investigate the interaction between the metallo-enzyme alcohol dehydrogenase (ADH) from yeast with bioflavonoid quercetin (QTN). Here, we have characterized the complex formation between QTN and Zn 2+ in aqueous solution and then examined the effect of such complex formation on the enzymatic activity of a zinc(II)-dependent enzyme alcohol dehydrogenase from yeast. We have observed an inhibition of enzymatic activity of ADH in presence of QTN. Enzyme inhibition kinetic experiments revealed QTN as a non-competitive inhibitor of yeast ADH. Perturbation of Circular dichroic (CD) spectrum of ADH in presence of QTN is observed due to the structural changes of ADH on complexation with the above flavonoid. Our results indicate a conformational change of ADH due to removal of Zn 2+ present in the enzyme by QTN. This was further established by molecular modeling study which shows that the flavonoid binds to the Zn 2+ ion which maintains the tertiary structure of the metallo-enzyme. So, QTN abstracts only half of the Zn 2+ ions present in the enzyme i.e. one Zn 2+ ion per monomer. From the present study, the structural alteration and loss of enzymatic activity of ADH are attributed to the complex formation between QTN and Zn 2+ . [ABSTRACT FROM AUTHOR]

Sowing time of soybean (Glycine max [L.] Merr.) often coincides with the early onset of rainy season. Germinating seedsencounter a transient to prolonged period of water-logging that causes anoxia (absence of oxygen) and hypoxia (insufficientoxygen) resulting in poor germination. This reduces crop stability and yield. One of the factors responsible for flood tolerance isactivity of alcohol dehydrogenase (ADH) during flood. The effect of ADH activity during flooding and difference in floodtolerance level were investigated using two soybean cultivars, Peking and Tamahomare, and their F9 recombinant inbred lines(RILs). Tamahomare showed higher ADH activity than Peking. There was a great variation in ADH activity among the RILs.QTL analysis detected five QTLs for ADH activity (qAas1-5) on five linkage groups, LG_A2, D1a, F, K and L. The QTL qAas4was close to a QTL for shoot damage and conductivity of germinating seeds after flooding treatment.

Flavour is a key quality attribute of apples defined by volatile aroma compounds. Biosynthesis of aroma compounds involves metabolic pathways in which the main precursors are fatty and amino acids, and the main products are aldehydes, alcohols and esters. Some enzymes are crucial in the production of volatile compounds, such as lipoxygenase, alcohol dehydrogenase, and alcohol acyltransferase. Composition and concentration of volatiles in apples may be altered by pre- and postharvest factors that cause a decline in apple flavour. Addition of biosynthetic precursors of volatile compounds may be a strategy to promote aroma production in apples. The present manuscript compiles information regarding the biosynthesis of volatile aroma compounds, including metabolic pathways, enzymes and substrates involved, factors that may affect their production and also includes a wide number of studies focused on the addition of biosynthetic precursors in their production. [ABSTRACT FROM AUTHOR]

Alcohol is eliminated from the body by various metabolic mechanisms. The primary enzymes in such mechanism involved are alcohol dehydrogenase, aldehyde dehydrogenase, cytochrome P450 2E1, and catalase. Variations in the genes for these enzymes have been found to influence alcohol consumption. The consequences of alcohol metabolism include oxygen deficits (i.e., hypoxia) in the liver, resulting in the formation of harmful compounds (i.e., adducts) and highly reactive oxygen-containing molecules (i.e., reactive oxygen species) that can damage cell components. Approximately, worldwide 3.6 % of cancers derive from chronic alcohol drinking, including those of the upper aerodigestive tract, the liver, the colorectum and the breast. Although the mechanisms for alcohol-associated carcinogenesis are not completely understood, recent findings have focused on acetaldehyde, the first and most toxic ethanol metabolite, as a cancer-causing agent. Alcohol-related carcinogenesis may aggravate due to other factors such as smoking and being triggered by genetic susceptibility. Besides, the role of genetic polymorphisms of the alcohol-metabolizing enzymes could not be ruled out. [ABSTRACT FROM AUTHOR]

A comparison on the larvicidial activity of four commercial bacterial and viral bioagents, Profect®, Virotecto®, Viroset® and Protecto® were evaluated on the 2nd and 4th larval instars of Spodoptera littoralis (Biosd.). The LC50 values showed 1.35, 1.52, 1.57 and 1.61 mg/ ml against 2nd instar larvae, respectively. On the other hand, the LC50 values recorded 2.03, 2.5, 2.72 and 3.01 mg/L. on 4th instar larvae of S. littoralis using the above mentioned commercial bioagent products, respectively. The effect on four isozymes, i.e., ,  esterase (EST), glutamate oxaloacetate transaminase (GOT), malate dehydrogenase (MDH) and alcohol dehydrogenase (ADH) were evaluated. The obtained results indicated differences in the activity of the isozymes in treated 4th instar larvae as compared to untreated larvae.

Molecular docking of substrates is more challenging compared to inhibitors as the reaction mechanism has to be considered. This becomes more pronounced for zinc-dependent enzymes since the coordination state of the catalytic zinc ion is of greater importance. In order to develop a predictive substrate docking protocol, we have performed molecular docking studies of diketone substrates using the catalytic state of carbonyl reductase 2 from Candida parapsilosis (CPCR2). Different docking protocols using two docking methods (AutoDock Vina and AutoDock4.2) with two different sets of atomic charges (AM1-BCC and HF-RESP) for catalytic zinc environment and substrates as well as two sets of vdW parameters for zinc ion were examined. We have selected the catalytic binding pose of each substrate by applying mechanism based distance criteria. To compare the performance of the docking protocols, the correlation plots for the binding energies of these catalytic poses were obtained against experimental V values of the 11 diketone substrates for CPCR2. The best correlation of 0.73 was achieved with AutoDock4.2 while treating catalytic zinc ion in optimized non-bonded (NB) state with +1.01 charge on the zinc ion, compared to 0.36 in non-bonded (+2.00 charge on the zinc ion) state. These results indicate the importance of catalytic constraints and charge parameterization of catalytic zinc environment for the prediction of substrate activity in zinc-dependent enzymes by molecular docking. The developed predictive docking protocol described here is in principle generally applicable for the efficient in silico substrate spectra characterization of zinc-dependent ADH. Graphical Abstract: [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

Objective: Identify and characterize polymorphisms of genes ADH2, ADH3, ALDH2 and CYP2E1 in a Colombian population residing in the city of Bogotá and determine its possible relationship to the alcoholism. Methods: ADH2, ADH3, ALDH2, and CYP2E1 genotypes a population of 148 individuals with non-problematic alcohol and 65 individuals with alcoholism were determined with TaqMan probes and PCR-RFLP. DNA was obtained from peripheral blood white cells. Results: Significant difference was found in family history of alcoholism and use of other psychoactive substances to compare alcoholics with controls. When allelic frequencies for each category (gender) were considered, frequency of A2 allele carriers in ADH2 was found higher in male patients than controls. In women, the relative frequency for c1 allele in CYP2E1 was lower in controls than alcoholics. The ALDH2 locus is monomorphic. No significant differences in allele distributions of the loci examined to compare two populations were observed, however when stratifying the same trend was found that these differences tended to be significant. Conclusions: This study allows us to conclude the positive association between family history of alcoholism and alcoholism suggesting that there is a favourable hereditary predisposition. Since substance dependence requires interaction of multiple genes, the combination of genotypes ADH2*2, CYP2E1*1 combined with genotype homozygous ALDH2*1 found in this study could be leading to the population to a potential risk to alcoholism. [ABSTRACT FROM AUTHOR]

It is now about half a century since molecular research on alcohol dehydrogenase (ADH), short-chain dehydrogenase/reductase (SDR) and medium-chain dehydrogenase/reductase (MDR) started. During this time, at least four stages of research can be distinguished, which led to many ADH, SDR and MDR structures from which their origins could be traced. An introductory summary of these stages is given, followed by a current update on the now known structures, including the present pattern of mammalian MDR–ADH enzymes into six classes and their evolutionary relationships. In spite of the wide spread in evolutionary changes from the “constant” class III to the more “variable” other classes, the change in class V (only confirmed as a transcript in humans) and class VI (absent in humans) are also restricted. Such spread in variability is visible also in other dehydrogenases, but not always so restricted in other co-evolving proteins we have studied. Finally, the shift in era of present ADH research is highlighted, as well as levels of likely future continuation. [ABSTRACT FROM AUTHOR]

Previous studies have reported that aspirin significantly reduced the first-pass metabolism (FPM) of ethanol in humans thereby increasing adverse effects of alcohol. The underlying causes, however, remain poorly understood. Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH), principal enzymes responsible for metabolism of ethanol, are complex enzyme families that exhibit functional polymorphisms among ethnic groups and distinct tissue distributions. We investigated the inhibition profiles by aspirin and its major metabolite salicylate of ethanol oxidation by recombinant human ADH1A, ADH1B1, ADH1B2, ADH1B3, ADH1C1, ADH1C2, ADH2, and ADH4, and acetaldehyde oxidation by ALDH1A1 and ALDH2, at pH 7.5 and 0.5 mM NAD + . Competitive inhibition pattern was found to be a predominant type among the ADHs and ALDHs studied, although noncompetitive and uncompetitive inhibitions were also detected in a few cases. The inhibition constants of salicylate for the ADHs and ALDHs were considerably lower than that of aspirin with the exception of ADH1A that can be ascribed to a substitution of Ala-93 at the bottom of substrate pocket as revealed by molecular docking experiments. Kinetic inhibition equation-based simulations show at higher therapeutic levels of blood plasma salicylate (1.5 mM) that the decrease of activities at 2–10 mM ethanol for ADH1A/ADH2 and ADH1B2/ADH1B3 are predicted to be 75–86% and 31–52%, respectively, and that the activity decline for ALDH1A1 and ALDH2 at 10–50 μM acetaldehyde to be 62–73%. Our findings suggest that salicylate may substantially inhibit hepatic FPM of alcohol at both the ADH and ALDH steps when concurrent intaking aspirin. [ABSTRACT FROM AUTHOR]

Abstract: In several animal species including humans, the acute administration of low doses of alcohol increases motor activity. Different theories have postulated that alcohol-induced hyperactivity is causally related to alcoholism. Moreover, a common biological mechanism in the mesolimbic dopamine system has been proposed to mediate the stimulant and motivational effects of alcohol. Numerous studies have examined whether alcohol-induced hyperactivity is related to alcoholism using a great variety of animal models and several animal species. However, there is no review that has summarized this extensive literature. In this article, we present the various experimental models that have been used to study the relationship between the stimulant and motivational effects of alcohol in rodents and primates. Furthermore, we discuss whether the theories hypothesizing a causal link between alcohol-induced hyperactivity and alcoholism are supported by published results. The reviewed findings indicate that animal species that are stimulated by alcohol also exhibit alcohol preference. Additionally, the role of dopamine in alcohol-induced hyperactivity is well established since blocking dopaminergic activity suppresses the stimulant effects of alcohol. However, dopamine transmission plays a much more complex function in the motivational properties of alcohol and the neuronal mechanisms involved in alcohol stimulation and reward are distinct. Overall, the current review provides mixed support for theories suggesting that the stimulant effects of alcohol are related to alcoholism and highlights the importance of animal models as a way to gain insight into alcoholism. [Copyright &y& Elsevier]

Antifreeze proteins are a class of polypeptides produced by certain animals, plants, fungi and bacteria that permit their survival under the subzero environments. Ammopiptanthus nanus is the unique evergreen broadleaf bush endemic to the Mid-Asia deserts. It survives at the west edge of the Tarim Basin from the disappearance of the ancient Mediterranean in the Tertiary Period. Its distribution region is characterized by the arid climate and extreme temperatures, where the extreme temperatures range from −30°C to 40°C. In the present study, the antifreeze protein gene AnAFP of A. nanus was used to transform Escherichia coli and tobacco, after bioinformatics analysis for its possible function. The transformed E. coli strain expressed the heterologous AnAFP gene under the induction of isopropyl β-D-thiogalactopyranoside, and demonstrated significant enhancement of cold tolerance. The transformed tobacco lines expressed the heterologous AnAFP gene in response to cold stress, and showed a less change of relative electrical conductivity under cold stress, and a less wilting phenotype after 16h of −3°C cold stress and thawing for 1h than the untransformed wild-type plants. All these results imply the potential value of the AnAFP gene to be used in genetic modification of commercially important crops for improvement of cold tolerance. [ABSTRACT FROM AUTHOR]

Highlights: [•] Methylglyoxal dehydrogenase activity is observed only in the glutathione-depleted cells. [•] The first report on methylglyoxal-oxidizing alcohol dehydrogenase in microorganisms. [•] Candida alcohol dehydrogenase is involved in methylglyoxal oxidation and reduction. [•] Accumulated methylglyoxal inhibits cell growth in alcohol dehydrogenase disruptants. [•] Alcohol dehydrogenase regulates cell cycle by catalyzing methylglyoxal in C. albicans. [ABSTRACT FROM AUTHOR]

Abstract: Objective: This study investigated the effects of scopoletin on alcohol-induced hepatic lipid accumulation in diet-induced obese mice and its mechanism. Material/Methods: Alcohol (25% v/v, 5g/kg body weight) was orally administered once a day for 6 weeks to mice fed with a high-fat diet (35%kcal) with or without scopoletin (0.05%, wt/wt). Results: Scopoletin reduced plasma acetaldehyde, fatty acid, total cholesterol, triglyceride and insulin levels, hepatic lipid and droplets and fasting blood glucose levels that were increased by alcohol. Scopoletin significantly activated hepatic AMPK and inhibited ACC and SREBP-1c and the activities of lipogenic enzymes, such as FAS, PAP and G6PD compared to the alcohol control group. Moreover, scopoletin significantly inhibited hepatic CYP2E1 activity and protein levels but elevated the activities of SOD, CAT, GSH-Px and GST and the levels of GSH compared to the alcohol control group. The hepatic lipid peroxide level was significantly lowered by scopoletin supplementation in alcohol-administered obese mice. Conclusions: Taken together, these results suggested that scopoletin can ameliorate alcohol-induced hepatic lipid accumulation by modulating AMPK–SREBP pathway-mediated lipogenesis in mice fed a high-fat diet. [Copyright &y& Elsevier]

Highlights: [•] A short-chain dehydrogenase/reductase (SM_SDR) from Serratia marcescens. [•] SM_SDR prefers NADPH as cofactor. [•] E. coli BL21 (DE3) expressing SM_SDR can produce (R)-PE from HPMAE. [•] Fructose is the most effective carbon source for biocatalysis. [ABSTRACT FROM AUTHOR]

Abstract: Alcohol dehydrogenase is a critical enzyme in the metabolism of alcohol. Expression of three alleles at the ADH1B locus results in enzymes that differ in turnover rate and affinity for alcohol. The ADH1B*3 allele, which appears to be unique to individuals of African descent, is associated with more rapid alcohol metabolism than the more prevalent ADH1B*1 allele. It has been previously demonstrated that the presence of at least one maternal ADH1B*3 allele confers a protective effect against alcohol teratogenicity in infants and children. This study was conducted to determine whether the presence of the ADH1B*3 allele in the mother or child continues to be protective in alcohol-exposed individuals during adolescence. 186 adolescents and 167 mothers participating in a 14-year follow-up of the Detroit Longitudinal Cohort were genotyped for ADH1B alleles. Behavioral reports were obtained from classroom teachers. Frequencies of the ADH1B*3 allele were 17.6% in the mothers and 21.0% in the adolescents, which are consistent with the 15–20% expected for African Americans. Prenatal alcohol exposure was associated with increased attention problems and externalizing behaviors in adolescents born to mothers with two ADH1B*1 alleles but not in those whose mothers had at least one ADH1B*3 allele. A similar pattern was seen in relation to the presence or absence of an ADH1B*3 allele in the adolescent, which may have reflected the presence/absence of the maternal variant. This study is the first to demonstrate that the protective effects of the maternal ADH1B*3 allele continue to be evident during adolescence. These persistent individual differences in vulnerability of offspring to the behavioral effects of fetal alcohol exposure are likely attributable to more rapid metabolism of alcohol that the ADH1B*3 variant confers on the mother, leading to a reduction of the peak blood alcohol concentration to which the fetus is exposed during each drinking episode. [Copyright &y& Elsevier]

Abstract: Calorie restriction (CR) is one of the most effective anti-aging interventions in mammals. A modern theory suggests that aging results from a decline in detoxification capabilities and thus accumulation of damaged macromolecules. The present study aimed to determine how short-term CR alters mRNA profiles of genes that encode metabolism and detoxification machinery in the liver. Male C57BL/6 mice were fed CR (0, 15, 30, or 40%) diets for one month, followed by mRNA quantification of 98 xenobiotic processing genes (XPGs) in the liver, including 7 uptake transporters, 39 phase-I enzymes, 37 phase-II enzymes, 10 efflux transporters, and 5 transcription factors. In general, 15% CR did not alter mRNAs of most XPGs, whereas 30 and 40% CR altered over half of the XPGs (32 increased and 29 decreased). CR up-regulated some phase-I enzymes (fold increase), such as Cyp4a14 (12), Por (2.3), Nqo1 (1.4), Fmo2 (5.4), and Fmo3 (346), and numerous number of phase-II enzymes, such as Sult1a1 (1.2), Sult1d1 (2.0), Sult1e1 (33), Sult3a1 (2.2), Gsta4 (1.3), Gstm2 (1.3), Gstm3 (1.7), and Mgst3 (2.2). CR feminized the mRNA profiles of 32 XPGs in livers of male mice. For instance, CR decreased the male-predominantly expressed Oatp1a1 (97%) and increased the female-predominantly expressed Oatp1a4 (11). In conclusion, short-term CR alters the mRNA levels of over half of the 98 XPGs quantified in livers of male mice, and over half of these alterations appear to be due to feminization of the liver. [Copyright &y& Elsevier]

Abstract: Background: To establish an infection in the vagina, Trichomonas vaginalis must adapt to various environmental cues for survival and further replication. Nutrient competition by lactobacilli, the major normal vaginal flora, is one of the mechanisms to limit the growth of other microorganisms. Additionally, lactobacilli produce H2O2 that can reduce the genital infections caused by other pathogens. Thus, the ability to overcome the metabolic stresses, such as glucose restriction (GR), as well as the oxidative stresses, is critical for T. vaginalis to establish an infection. Methods: To gain insights into the molecular mechanisms of adaptation to GR, we utilized next-generation RNA sequencing (RNA-seq) to quantify the gene expression changes upon GR. Autophagy, a cytoprotective response to starvation, was monitored by using autophagy-specific staining, autophagy inhibition assay, and co-localization of autophagosomes with lysosomes. Results: We demonstrated that GR promotes the survival of T. vaginalis. Besides, GR-cultivated cells exhibit higher H2O2 resistance. Our RNA-seq data revealed that genes involved in general energy metabolism were downregulated, whereas genes encoding glutamate metabolism-related aminotransferases were strikingly upregulated under GR. Furthermore, autophagy was first identified and characterized in T. vaginalis under GR. Conclusions: These data suggest that GR induces a metabolic reprogramming, enhancing antioxidant ability and autophagy for cellular homeostasis to maintain survival. General significance: Our work not only led to significant advances in understanding the transcriptional changes in response to GR but also provided possible strategies elicited by GR for T. vaginalis to adapt to the vaginal microenvironment. [Copyright &y& Elsevier]

Highlights: [•] FXR activity was impaired by chronic ethanol ingestion in a murine model of ALD. [•] Activation of FXR attenuated alcohol-induced liver injury and steatosis. [•] Activation of FXR attenuated cholestasis and oxidative stress in mouse liver. [ABSTRACT FROM AUTHOR]

Abstract: Ethnopharmacologic relevance: The Chunggan extract (CGX) is a traditional herbal formula prescribed for patients suffering from various liver diseases, including alcoholic liver disease, in which the mechanism of CGX action remains unclear. This study aimed to investigate the anti-hepatic fibrosis effects of CGX and its underlying mechanisms in alcohol-induced rat livers. Materials and methods: To elucidate the mechanism of action of CGX, we evaluated gene expression profiles in the livers of rats treated with 30% alcohol and anti-fibrotic doses of CGX of 100 and 200mg/kg/day at 1 day, and 2 and 4 weeks using microarrays. The mRNA and protein expression levels of vascular endothelial growth factor (VEGF), one of the candidate genes selected in this study, in alcohol-induced rat livers were measured by real-time PCR and enzyme-linked immunosorbent assays, respectively. Results: We identified 4128 genes as differentially expressed by at least twofold between alcohol-only– and alcohol–CGX–fed rats at various doses and time points, compared to naïve control animals. Twenty-three of these genes were associated with liver fibrosis and oxidative stress based on the GeneCards database, resulting in p<0.05 by ANOVA between the alcohol-only and alcohol–CGX groups. Especially, Vegf was decreased in CGX 200mg/kg/day-fed rat livers at all time points evaluated, and mRNA and protein levels at the 4-week time point were validated. Conclusion: These gene expression profiles provide a better understanding of the mechanisms underlying the anti-fibrotic effects of CGX. Suppression of VEGF may play a critical role in anti-fibrotic action of CGX in alcoholic liver injury. [Copyright &y& Elsevier]

Highlights: [•] Gene expression profile was examined in RhE model. [•] Five cellular stress related genes were significantly upregulated by DNFB and OXA. [•] Predictive performance of ATF3 gene displayed 100% accuracy to animal testing. [•] Tested four pre-/pro-haptens were correctly predicted by ATF3 and DNAJB4 genes. [•] A RhE-based assay could detect skin sensitizing potential of chemicals. [Copyright &y& Elsevier]

Abstract: The thymus provides the microenvironment in which thymocytes develop into mature T-cells, and interactions with thymic stromal cells are thought to provide the necessary signals for thymocyte maturation. Recognition of self-MHC by T-cells is a basic requirement for mature T-cell functions, and those thymocytes that do not recognize or respond too strongly to the peptide-loaded self-MHC molecules found in the thymus undergo apoptosis. As a result, 95% of the thymocytes produced will die and be subsequently cleared by macrophages. This review describes a complex crosstalk between developing thymocytes and engulfing macrophages which is mediated by retinoids produced by engulfing macrophages. The interaction results in the harmonization of the rate of cell death of dying double positive cells with their clearance and replacement, and in promotion of the differentiation of the selected cells in the thymus. [Copyright &y& Elsevier]

Abstract: Metabolic engineering has been playing important roles in developing high performance microorganisms capable of producing various chemicals and materials from renewable biomass in a sustainable manner. Synthetic and systems biology are also contributing significantly to the creation of novel pathways and the whole cell-wide optimization of metabolic performance, respectively. In order to expand the spectrum of chemicals that can be produced biotechnologically, it is necessary to broaden the metabolic capacities of microorganisms. Expanding the metabolic pathways for biosynthesizing the target chemicals requires not only the enumeration of a series of known enzymes, but also the identification of biochemical gaps whose corresponding enzymes might not actually exist in nature; this issue is the focus of this paper. First, pathway prediction tools, effectively combining reactions that lead to the production of a target chemical, are analyzed in terms of logics representing chemical information, and designing and ranking the proposed metabolic pathways. Then, several approaches for potentially filling in the gaps of the novel metabolic pathway are suggested along with relevant examples, including the use of promiscuous enzymes that flexibly utilize different substrates, design of novel enzymes for non-natural reactions, and exploration of hypothetical proteins. Finally, strain optimization by systems metabolic engineering in the context of novel metabolic pathways constructed is briefly described. It is hoped that this review paper will provide logical ways of efficiently utilizing ‘big’ biological data to design and develop novel metabolic pathways for the production of various bulk chemicals that are currently produced from fossil resources. [Copyright &y& Elsevier]

Highlights: [•] A PCR strategy was used to isolate a full-length CgADH cDNA from chrysanthemum. [•] The hypoxic induction of CgADH can be inhibited by ethylene in C. zawadskii. [•] Ethylene is required for the adaptation of chrysanthemum to waterlogged conditions. [Copyright &y& Elsevier]