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MBio Jan 2019Butanol production by is accompanied by coproduction of acetone and ethanol, which reduces the yield of butanol and increases the production cost. Here, we report...
Butanol production by is accompanied by coproduction of acetone and ethanol, which reduces the yield of butanol and increases the production cost. Here, we report development of several clostridial aldehyde/alcohol dehydrogenase (AAD) variants showing increased butanol selectivity by a series of design and analysis procedures, including random mutagenesis, substrate specificity feature analysis, and structure-based butanol selectivity design. The butanol/ethanol ratios (B/E ratios) were dramatically increased to 17.47 and 15.91 g butanol/g ethanol for AAD and AAD, respectively, which are 5.8-fold and 5.3-fold higher than the ratios obtained with the wild-type AAD. The much-increased B/E ratio obtained was due to the dramatic reduction in ethanol production (0.59 ± 0.01 g/liter) that resulted from engineering the substrate binding chamber and the active site of AAD. This protein design strategy can be applied generally for engineering enzymes to alter substrate selectivity. Renewable biofuel represents one of the answers to solving the energy crisis and climate change problems. Butanol produced naturally by clostridia has superior liquid fuel characteristics and thus has the potential to replace gasoline. Due to the lack of efficient genetic manipulation tools, however, clostridial strain improvement has been slower than improvement of other microorganisms. Furthermore, fermentation coproducing various by-products requires costly downstream processing for butanol purification. Here, we report the results of enzyme engineering of aldehyde/alcohol dehydrogenase (AAD) to increase butanol selectivity. A metabolically engineered strain expressing the engineered aldehyde/alcohol dehydrogenase gene was capable of producing butanol at a high level of selectivity.
Topics: Acetone; Alcohol Dehydrogenase; Aldehyde Dehydrogenase; Butanols; Catalytic Domain; Clostridium acetobutylicum; Ethanol; Fermentation; Metabolic Engineering; Molecular Dynamics Simulation; Mutagenesis, Site-Directed
PubMed: 30670620
DOI: 10.1128/mBio.02683-18 -
Biochemistry and Molecular Biology... Jan 2022Project-based (research-driven) laboratory courses stimulate student involvement, improve critical thinking and initiate cooperative learning. To this end, a 7-week...
Project-based (research-driven) laboratory courses stimulate student involvement, improve critical thinking and initiate cooperative learning. To this end, a 7-week laboratory project was designed for a sophomore cell biology course to reinforce the fundamental relationship between genotype and phenotype using yeast alcohol dehydrogenase I (ADH1). Working in pairs, students used site-directed mutagenesis to create a H44R mutation in the ADH1 gene sequence inserted into a YEp13 shuttle vector. These plasmids were propagated in E. coli, sequenced, and reintroduced into a yeast strain expressing no ADH1 activity. The growth patterns on selective media were determined. As the mutation allows for growth in the presence of allyl alcohol, students make the connection between DNA sequence and protein function. Student performance was assessed with pre- and post-tests, with improvement observed across all learning objectives. In addition to meeting the learning outcomes, 98% of the students thought that this experience allowed them to see how the scientific process can encompass multiple techniques to answer a single question. Eighty-four percent of the students thought that this experience was more engaging than other lab experiences they have had. Our multi-week laboratory examining the phenotypic changes in yeast alcohol metabolism successfully developed students' understanding of the scientific process, knowledge of molecular techniques and the relationship of gene sequence to protein function in an engaging manner.
Topics: Alcohol Dehydrogenase; Escherichia coli; Genotype; Humans; Phenotype; Proteins; Saccharomyces cerevisiae
PubMed: 34878214
DOI: 10.1002/bmb.21594 -
Molecules (Basel, Switzerland) Jul 2023A biomimetic mineralization method was used in the facile and rapid preparation of nanoflowers for immobilizing alcohol dehydrogenase (ADH). The method mainly uses ADH...
A biomimetic mineralization method was used in the facile and rapid preparation of nanoflowers for immobilizing alcohol dehydrogenase (ADH). The method mainly uses ADH as an organic component and zinc phosphate as an inorganic component to prepare flower-like ADH/Zn(PO) organic-inorganic hybrid nanoflowers (HNFs) with the high specific surface area through a self-assembly process. The synthesis conditions of the ADH HNFs were optimized and its morphology was characterized. Under the optimum enzymatic reaction conditions, the Michaelis-Menten constant () of ADH HNFs (β-NAD as substrate) was measured to be 3.54 mM, and the half-maximal inhibitory concentration (IC) of the positive control ranitidine (0.2-0.8 mM) was determined to be 0.49 mM. Subsequently, the inhibitory activity of natural medicine Pursh and nine small-molecule compounds on ADH was evaluated using ADH HNFs. The inhibition percentage of the aqueous extract of is 57.9%. The vanillic acid, protocatechuic acid, gallic acid, and naringenin have obvious inhibitory effects on ADH, and their percentages of inhibition are 55.1%, 68.3%, 61.9%, and 75.5%, respectively. Moreover, molecular docking analysis was applied to explore the binding modes and sites of the four most active small-molecule compounds to ADH. The results of this study can broaden the application of immobilized enzymes through biomimetic mineralization, and provide a reference for the discovery of ADH inhibitors from natural products.
Topics: Alcohol Dehydrogenase; Nanostructures; Biomimetics; Molecular Docking Simulation
PubMed: 37513303
DOI: 10.3390/molecules28145429 -
British Journal of Cancer Feb 2023Increases in IL-6 by cancer-associated fibroblasts (CAFs) contribute to colon cancer progression, but the mechanisms involved in the increase of this tumor-promoting...
BACKGROUND
Increases in IL-6 by cancer-associated fibroblasts (CAFs) contribute to colon cancer progression, but the mechanisms involved in the increase of this tumor-promoting cytokine are unknown. The aim of this study was to identify novel targets involved in the dysregulation of IL-6 expression by CAFs in colon cancer.
METHODS
Colonic normal (N), hyperplastic, tubular adenoma, adenocarcinoma tissues, and tissue-derived myo-/fibroblasts (MFs) were used in these studies.
RESULTS
Transcriptomic analysis demonstrated a striking decrease in alcohol dehydrogenase 1B (ADH1B) expression, a gene potentially involved in IL-6 dysregulation in CAFs. ADH1B expression was downregulated in approximately 50% of studied tubular adenomas and all T1-4 colon tumors, but not in hyperplastic polyps. ADH1B metabolizes alcohols, including retinol (RO), and is involved in the generation of all-trans retinoic acid (atRA). LPS-induced IL-6 production was inhibited by either RO or its byproduct atRA in N-MFs, but only atRA was effective in CAFs. Silencing ADH1B in N-MFs significantly upregulated LPS-induced IL-6 similar to those observed in CAFs and lead to the loss of RO inhibitory effect on inducible IL-6 expression.
CONCLUSION
Our data identify ADH1B as a novel potential mesenchymal tumor suppressor, which plays a critical role in ADH1B/retinoid-mediated regulation of tumor-promoting IL-6.
Topics: Humans; Alcohol Dehydrogenase; Cancer-Associated Fibroblasts; Colonic Neoplasms; Fibroblasts; Interleukin-6; Lipopolysaccharides; Tretinoin; Vitamin A
PubMed: 36482184
DOI: 10.1038/s41416-022-02066-0 -
PloS One 2021Observational studies suggest alcohol use promotes the development of some adverse cardiometabolic traits but protects against others including outcomes related to...
Observational studies suggest alcohol use promotes the development of some adverse cardiometabolic traits but protects against others including outcomes related to coronary artery disease. We used Mendelian randomization (MR) to explore causal relationships between the degree of alcohol consumption and several cardiometabolic traits in the UK Biobank. Using the well-established ADH1B Arg47His variant (rs1229984) and up to 24 additional SNPs recently found to be associated with alcohol consumption in an independent dataset as instruments, we conducted two-stage least squares and inverse weighted variance MR analyses, both as one-sample analyses in the UK Biobank and as two-sample analyses in external consortia. In the UK Biobank inverse variance weighted analyses, we found that one additional drink of alcohol per day was positively associated with systolic blood pressure (beta = 2.65 mmHg [1.40, 3.89]), hemorrhagic stroke (OR = 2.25 [1.41, 3.60]), and atrial fibrillation (OR = 1.26 [1.07, 1.48]), which were replicated in multivariable analyses. Alcohol was also associated with all cardiovascular disease and all-cause death. A positive association with myocardial infarction did not replicate in multivariable analysis, with suggestive mediation through blood pressure; similarly, a positive association between alcohol use with type 2 diabetes was mitigated by BMI in multivariable analysis. Findings were generally null in replication with two-sample analyses. Alcohol was not protective for any disease outcome with any analysis method, dataset, or strata. Stratifications by sex and smoking in the UK Biobank revealed higher point estimates of risk for several outcomes for men and mixed results for smoking strata, but no statistically significant heterogeneity. Our results are consistent with an overall harmful and/or null effect of alcohol on cardiometabolic health at all levels of use and suggest that even moderate alcohol use should not be promoted as a part of a healthy diet and lifestyle.
Topics: Adult; Aged; Alcohol Dehydrogenase; Alcohol Drinking; Atrial Fibrillation; Biological Specimen Banks; Blood Pressure; Body Mass Index; Cardiovascular Diseases; Diabetes Mellitus, Type 2; Female; Hemorrhagic Stroke; Humans; Male; Mendelian Randomization Analysis; Middle Aged; Polymorphism, Single Nucleotide; Prospective Studies; United Kingdom
PubMed: 34379647
DOI: 10.1371/journal.pone.0255801 -
Redox Biology May 2024Elevated fasting ethanol levels in peripheral blood frequently found in metabolic dysfunction-associated steatohepatitis (MASLD) patients even in the absence of alcohol...
Elevated fasting ethanol levels in peripheral blood frequently found in metabolic dysfunction-associated steatohepatitis (MASLD) patients even in the absence of alcohol consumption are discussed to contribute to disease development. To test the hypothesis that besides an enhanced gastrointestinal synthesis a diminished alcohol elimination through alcohol dehydrogenase (ADH) may also be critical herein, we determined fasting ethanol levels and ADH activity in livers and blood of MASLD patients and in wild-type ± anti-TNFα antibody (infliximab) treated and TNFα mice fed a MASLD-inducing diet. Blood ethanol levels were significantly higher in patients and wild-type mice with MASLD while relative ADH activity in blood and liver tissue was significantly lower compared to controls. Both alterations were significantly attenuated in MASLD diet-fed TNFα mice and wild-type mice treated with infliximab. Moreover, alcohol elimination was significantly impaired in mice with MASLD. In in vitro models, TNFα but not IL-1β or IL-6 significantly decreased ADH activity. Our data suggest that elevated ethanol levels in MASLD patients are related to TNFα-dependent impairments of ADH activity.
Topics: Mice; Humans; Animals; Alcohol Dehydrogenase; Tumor Necrosis Factor-alpha; Infliximab; Ethanol; Fatty Liver; Alcohol Drinking
PubMed: 38493749
DOI: 10.1016/j.redox.2024.103121 -
Behavioural Neurology 2021Aldehyde dehydrogenase 2 () polymorphisms are related to both stroke risk and alcohol consumption. However, the influence of polymorphisms and alcohol consumption on...
Aldehyde dehydrogenase 2 () polymorphisms are related to both stroke risk and alcohol consumption. However, the influence of polymorphisms and alcohol consumption on cognitive impairment after ischemic stroke remains unknown, as do the possible mechanisms. We enrolled 180 Han Chinese ischemic stroke patients from four community health centers in Bengbu, China. Cognitive function was assessed using the Montreal Cognitive Assessment (MoCA), and two different MoCA cutoff scores were used to define cognitive impairment in ischemic stroke patients. The genotypes were determined using polymerase chain reaction and direct sequencing. To assess the associations of polymorphisms and alcohol consumption with cognitive impairment after ischemic stroke, we performed binary logistic regression analysis with odds ratios. We revealed that individuals with the wild-type genotype were more likely to have high MoCA scores than those with the mutant and heterozygous types ( = 0.034). In addition, using two MoCA cutoff scores, the percentage of moderate to excessive alcohol consumption in the cognitive impairment group was higher than that in the nonimpairment group ( = 0.001). The levels of 4-hydroxy-2-nonenal ( = 0.001) and swallowing function ( = 0.001) were also higher in the cognitive impairment group than in the nonimpairment group. Moreover, after adjusting for other potential risk factors, polymorphisms and alcohol consumption had a significant synergistic effect on cognitive impairment ( = 0.022). Specifically, the ∗ mutant allele and higher alcohol consumption were associated with cognitive impairment and swallowing ability after ischemic stroke. Targeting ALDH2 may be a useful biomarker for cognitive rehabilitation following ischemic stroke.
Topics: Alcohol Dehydrogenase; Alcohol Drinking; Aldehyde Dehydrogenase; Aldehyde Dehydrogenase, Mitochondrial; Brain Ischemia; China; Cognitive Dysfunction; Genotype; Humans; Ischemic Stroke; Stroke
PubMed: 34257742
DOI: 10.1155/2021/6696806 -
Genes Aug 2022In the present review, the main features involved in the susceptibility and progression of neurodegenerative disorders (NDDs) have been discussed, with the purpose of... (Review)
Review
In the present review, the main features involved in the susceptibility and progression of neurodegenerative disorders (NDDs) have been discussed, with the purpose of highlighting their potential application for promoting the management and treatment of patients with NDDs. In particular, the impact of genetic and epigenetic factors, nutrients, and lifestyle will be presented, with particular emphasis on Alzheimer's disease (AD) and Parkinson's disease (PD). Metabolism, dietary habits, physical exercise and microbiota are part of a complex network that is crucial for brain function and preservation. This complex equilibrium can be disrupted by genetic, epigenetic, and environmental factors causing perturbations in central nervous system homeostasis, contributing thereby to neuroinflammation and neurodegeneration. Diet and physical activity can directly act on epigenetic modifications, which, in turn, alter the expression of specific genes involved in NDDs onset and progression. On this subject, the introduction of nutrigenomics shed light on the main molecular players involved in the modulation of health and disease status. In particular, the review presents data concerning the impact of , and on the susceptibility and progression of NDDs (especially AD and PD) and how they may be exploited for developing precision medicine strategies for the disease treatment and management.
Topics: Alcohol Dehydrogenase; Alzheimer Disease; Cytochrome P-450 CYP1A2; Epigenesis, Genetic; Humans; Life Style; Methylenetetrahydrofolate Reductase (NADPH2); Neurodegenerative Diseases; Nutritional Status; Parkinson Disease
PubMed: 36011409
DOI: 10.3390/genes13081498 -
Science Advances Jan 2020High alcohol consumption is a risk factor for morbidity and mortality, yet few genetic loci have been robustly associated with alcohol intake. Here, we use U.K. Biobank...
High alcohol consumption is a risk factor for morbidity and mortality, yet few genetic loci have been robustly associated with alcohol intake. Here, we use U.K. Biobank ( = 125,249) and GERA ( = 47,967) datasets to determine genetic factors associated with extreme population-level alcohol consumption and examine the functional validity of outcomes using model organisms and in silico techniques. We identified six loci attaining genome-wide significant association with alcohol consumption after meta-analysis and meeting our criteria for replication: (lead SNP: rs1229984), (rs13130794), (rs144198753), (rs1260326), (rs13107325), and (rs11214609). A conserved role in phenotypic responses to alcohol was observed for all genetic targets available for investigation (, and ) in . Evidence of causal links to lung cancer, and shared genetic architecture with gout and hypertension was also found. These findings offer insight into genes, pathways, and relationships for disease risk associated with high alcohol consumption.
Topics: Alcohol Dehydrogenase; Alcohol Drinking; Alcoholism; Biomarkers; Female; Genetic Association Studies; Genetic Loci; Genetic Predisposition to Disease; Genome-Wide Association Study; Humans; Linkage Disequilibrium; Male; Phenotype; Polymorphism, Single Nucleotide; Quantitative Trait Loci; Signal Transduction
PubMed: 31998841
DOI: 10.1126/sciadv.aay5034 -
Applied and Environmental Microbiology Oct 2018Fructophilic lactic acid bacteria (FLAB) are a recently discovered group, consisting of a few and species. Because of their unique characteristics, including poor... (Review)
Review
Fructophilic lactic acid bacteria (FLAB) are a recently discovered group, consisting of a few and species. Because of their unique characteristics, including poor growth on glucose and preference of oxygen, they are regarded as "unconventional" lactic acid bacteria (LAB). Their unusual growth characteristics are due to an incomplete gene encoding a bifunctional alcohol/acetaldehyde dehydrogenase (). This results in the imbalance of NAD/NADH and the requirement of additional electron acceptors to metabolize glucose. Oxygen, fructose, and pyruvate are used as electron acceptors. FLAB have significantly fewer genes for carbohydrate metabolism than other LAB, especially due to the lack of complete phosphotransferase system (PTS) transporters. They have been isolated from fructose-rich environments, including flowers, fruits, fermented fruits, and the guts of insects that feed on plants rich in fructose, and are separated into two groups on the basis of their habitats. One group is associated with flowers, grapes, wines, and insects, and the second group is associated with ripe fruits and fruit fermentations. Species associated with insects may play a role in the health of their host and are regarded as suitable vectors for paratransgenesis in honey bees. Besides their impact on insect health, FLAB may be promising candidates for the promotion of human health. Further studies are required to explore their beneficial properties in animals and humans and their applications in the food industry.
Topics: Alcohol Dehydrogenase; Aldehyde Oxidoreductases; Animals; Bacterial Proteins; Bees; Fermentation; Flowers; Fructose; Fruit; Glucose; Insecta; Lactobacillales; Lactobacillus; Leuconostocaceae; Phylogeny; Wine
PubMed: 30054367
DOI: 10.1128/AEM.01290-18