-
Chembiochem : a European Journal of... Mar 2022In this study, we developed a new bienzymatic reaction to produce enantioenriched phenylethanols. In a first step, the recombinant, unspecific peroxygenase from Agrocybe...
In this study, we developed a new bienzymatic reaction to produce enantioenriched phenylethanols. In a first step, the recombinant, unspecific peroxygenase from Agrocybe aegerita (rAaeUPO) was used to oxidise ethylbenzene and its derivatives to the corresponding ketones (prochiral intermediates) followed by enantioselective reduction into the desired (R)- or (S)-phenylethanols using the (R)-selective alcohol dehydrogenase (ADH) from Lactobacillus kefir (LkADH) or the (S)-selective ADH from Rhodococcus ruber (ADH-A). In a one-pot two-step cascade, 11 ethylbenzene derivatives were converted into the corresponding chiral alcohols at acceptable yields and often excellent enantioselectivity.
Topics: Alcohol Dehydrogenase; Benzene Derivatives; Mixed Function Oxygenases; Oxidation-Reduction; Phenylethyl Alcohol; Stereoisomerism
PubMed: 35023279
DOI: 10.1002/cbic.202200017 -
Toxicological Sciences : An Official... Aug 2018Alcohol metabolism is a well-characterized biological process that is dominated by the alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) families....
Alcohol metabolism is a well-characterized biological process that is dominated by the alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) families. Nonalcoholic steatohepatitis (NASH) is the advanced inflammatory stage of nonalcoholic fatty liver disease (NAFLD) and is known to alter the metabolism and disposition of numerous drugs. The purpose of this study was to investigate the alterations in alcohol metabolism processes in response to human NASH progression. Expression and function of ADHs, ALDHs, and catalase were examined in normal, steatosis, NASH (fatty) and NASH (not fatty) human liver samples. ALDH4A1 mRNA was significantly decreased in both NASH groups, while no significant changes were observed in the mRNA levels of other alcohol-related enzymes. The protein levels of ADH1A, ADH1B, and ADH4 were each decreased in the NASH groups, which was consistent with a decreased overall ADH activity. The protein level of ALDH2 was significantly increased in both NASH groups, while ALDH1A1 and ALDH1B1 were only decreased in NASH (fatty) samples. ALDH activity represented by oxidation of acetaldehyde was decreased in the NASH (fatty) group. The protein level of catalase was decreased in both NASH groups, though activity was unchanged. Furthermore, the significant accumulation of 4-hydroxynonenal protein adduct in NASH indicated significant oxidative stress and a potential reduction in ALDH activity. Collectively, ADH and ALDH expression and function are profoundly altered in the progression of NASH, which may have a notable impact on ADH- and ALDH-associated cellular metabolism processes and lead to significant alterations in drug metabolism mediated by these enzymes.
Topics: Alcohol Dehydrogenase; Aldehyde Dehydrogenase; Disease Progression; Ethanol; Humans; Isoenzymes; Liver; Non-alcoholic Fatty Liver Disease; Polymorphism, Single Nucleotide; RNA, Messenger
PubMed: 29718361
DOI: 10.1093/toxsci/kfy106 -
Cellular and Molecular Life Sciences :... Dec 2008Zinc plays an important role in the structure and function of many enzymes, including alcohol dehydrogenases (ADHs) of the MDR type (mediumchain... (Review)
Review
Zinc plays an important role in the structure and function of many enzymes, including alcohol dehydrogenases (ADHs) of the MDR type (mediumchain dehydrogenases/reductases). Active site zinc participates in catalytic events, and structural site zinc maintains structural stability. MDR-types of ADHs have both of these zinc sites but with some variation in ligands and spacing. The catalytic zinc sites involve three residues with different spacings from two separate protein segments, while the structural zinc sites involve four residues and cover a local segment of the protein chain (Cys97-Cys111 in horse liver class I ADH). This review summarizes properties of both ADH zinc sites, and relates them to zinc sites of proteins in general. In addition, it highlights a separate study of zinc binding peptide variants of the horse liver ADH structural zinc site. The results show that zinc coordination of the free peptide differs markedly from that of the enzyme (one His / three Cys versus four Cys), suggesting that the protein zinc site is in an energetically strained conformation relative to that of the peptide. This finding is a characteristic of an entatic state, implying a functional nature for this zinc site.
Topics: Alcohol Dehydrogenase; Amino Acid Sequence; Animals; Catalysis; Catalytic Domain; Humans; Molecular Sequence Data; Multigene Family; Peptides; Zinc
PubMed: 19011745
DOI: 10.1007/s00018-008-8593-1 -
International Journal of Molecular... Sep 2021Most bacteria possess alcohol dehydrogenase (ADH) genes ( genes) to mitigate alcohol toxicity, but these genes have functions beyond alcohol degradation. Previous...
Most bacteria possess alcohol dehydrogenase (ADH) genes ( genes) to mitigate alcohol toxicity, but these genes have functions beyond alcohol degradation. Previous research has shown that ADH can modulate quorum sensing in , a rising opportunistic pathogen. However, the number and nature of genes in have not yet been fully characterized. We identified seven alcohol dehydrogenases (NAD-ADHs) from ATCC 19606, and examined the roles of three iron-containing ADHs, ADH3, ADH4, and ADH6. Marker-less mutation was used to generate , , and single, double, and triple mutants. Disrupted mutants failed to grow in ethanol-, 1-butanol-, or 1-propanol-containing mediums, and recombinant ADH4 exhibited strongest activity against ethanol. Stress resistance assays with inorganic and organic hydroperoxides showed that and were key to oxidative stress resistance. Virulence assays performed on the model organism revealed that mutants had comparable virulence to wild-type, while and mutants had reduced virulence. The results suggest that ADH4 is primarily involved in alcohol metabolism, while ADH3 and ADH6 are key to stress resistance and virulence. Further investigation into the roles of other ADHs in is warranted.
Topics: Acinetobacter baumannii; Alcohol Dehydrogenase; Amino Acid Sequence; Animals; Computer Simulation; Cytosol; Ethanol; Gene Expression Regulation, Bacterial; Genes, Bacterial; Homeostasis; Hydrogen-Ion Concentration; Iron; Melanins; Moths; Mutation; Oxidative Stress; Phylogeny; Stress, Physiological; Virulence
PubMed: 34576087
DOI: 10.3390/ijms22189921 -
BMC Genomics Mar 2014The alcohol dehydrogenase (ADH) gene family uniquely illustrates the concept of enzymogenesis. In vertebrates, tandem duplications gave rise to a multiplicity of forms...
BACKGROUND
The alcohol dehydrogenase (ADH) gene family uniquely illustrates the concept of enzymogenesis. In vertebrates, tandem duplications gave rise to a multiplicity of forms that have been classified in eight enzyme classes, according to primary structure and function. Some of these classes appear to be exclusive of particular organisms, such as the frog ADH8, a unique NADP+-dependent ADH enzyme. This work describes the ADH system of Xenopus, as a model organism, and explores the first amphibian and reptilian genomes released in order to contribute towards a better knowledge of the vertebrate ADH gene family.
RESULTS
Xenopus cDNA and genomic sequences along with expressed sequence tags (ESTs) were used in phylogenetic analyses and structure-function correlations of amphibian ADHs. Novel ADH sequences identified in the genomes of Anolis carolinensis (anole lizard) and Pelodiscus sinensis (turtle) were also included in these studies. Tissue and stage-specific libraries provided expression data, which has been supported by mRNA detection in Xenopus laevis tissues and regulatory elements in promoter regions. Exon-intron boundaries, position and orientation of ADH genes were deduced from the amphibian and reptilian genome assemblies, thus revealing syntenic regions and gene rearrangements with respect to the human genome. Our results reveal the high complexity of the ADH system in amphibians, with eleven genes, coding for seven enzyme classes in Xenopus tropicalis. Frogs possess the amphibian-specific ADH8 and the novel ADH1-derived forms ADH9 and ADH10. In addition, they exhibit ADH1, ADH2, ADH3 and ADH7, also present in reptiles and birds. Class-specific signatures have been assigned to ADH7, and ancestral ADH2 is predicted to be a mixed-class as the ostrich enzyme, structurally close to mammalian ADH2 but with class-I kinetic properties. Remarkably, many ADH1 and ADH7 forms are observed in the lizard, probably due to lineage-specific duplications. ADH4 is not present in amphibians and reptiles.
CONCLUSIONS
The study of the ancient forms of ADH2 and ADH7 sheds new light on the evolution of the vertebrate ADH system, whereas the special features showed by the novel forms point to the acquisition of new functions following the ADH gene family expansion which occurred in amphibians.
Topics: Alcohol Dehydrogenase; Amino Acid Sequence; Amphibians; Animals; Evolution, Molecular; Expressed Sequence Tags; Female; Gene Library; Genome; Molecular Sequence Data; Phylogeny; Promoter Regions, Genetic; Reptiles; Sequence Alignment; Xenopus
PubMed: 24649825
DOI: 10.1186/1471-2164-15-216 -
Communications Biology Jun 2020Aldehyde-alcohol dehydrogenase (AdhE) is an enzyme responsible for converting acetyl-CoA to ethanol via acetaldehyde using NADH. AdhE is composed of two catalytic...
Aldehyde-alcohol dehydrogenase (AdhE) is an enzyme responsible for converting acetyl-CoA to ethanol via acetaldehyde using NADH. AdhE is composed of two catalytic domains of aldehyde dehydrogenase (ALDH) and alcohol dehydrogenase (ADH), and forms a spirosome architecture critical for AdhE activity. Here, we present the atomic resolution (3.43 Å) cryo-EM structure of AdhE spirosomes in an extended conformation. The cryo-EM structure shows that AdhE spirosomes undergo a structural transition from compact to extended forms, which may result from cofactor binding. This transition leads to access to a substrate channel between ALDH and ADH active sites. Furthermore, prevention of this structural transition by crosslinking hampers the activity of AdhE, suggesting that the structural transition is important for AdhE activity. This work provides a mechanistic understanding of the regulation mechanisms of AdhE activity via structural transition, and a platform to modulate AdhE activity for developing antibiotics and for facilitating biofuel production.
Topics: Alcohol Dehydrogenase; Aldehyde Oxidoreductases; Aldehydes; Cryoelectron Microscopy; Crystallography, X-Ray; Escherichia coli; Escherichia coli Proteins; Ethanol; Models, Molecular; Organelles; Protein Conformation; Substrate Specificity
PubMed: 32523125
DOI: 10.1038/s42003-020-1030-1 -
Applied and Environmental Microbiology Sep 2017Proteins with high catalytic efficiency and selectivity under mild conditions have long been appreciated by industrial and medicinal fields. These proteins, which are...
Proteins with high catalytic efficiency and selectivity under mild conditions have long been appreciated by industrial and medicinal fields. These proteins, which are commonly multimeric, often possess low stability, impeding wider application. Currently, strategies to improve the stability of multimeric proteins concentrate on enhancing the interaction at internal interface of the subunits. In this report, we confirmed that the largely underestimated subunit terminal ends are as significant as the internal interface for protein stability. By connecting both the terminal ends and internal interface of subunits, the tetrameric alcohol dehydrogenase (ADH) protein can been cyclized into a rigid form with significantly improved thermostability and resilience. The improvement in the temperature at which enzyme activity is reduced to 50% after a 15-min heat treatment () and melting temperature ( ) of the modified protein was 18°C and 23.3°C, respectively, which is superior to the results achieved by normal protein engineering. Our study provided a novel strategy to effectively improve the stability of multimeric proteins, which is suitable not only for the short-chain dehydrogenase/reductase (SDR) family but also other classes of proteins with close terminal ends. Industrially interesting proteins are generally multimeric proteins; however, their applications are often restricted due to low stability caused by the natural tendency of subunit disassociation. Current approaches targeting this problem mainly focus on enhancing the internal interfaces of the subunits to avoid their disassociation. In this study, we identified and confirmed the external interface to be significant for improving the stability of multimeric proteins. By connecting the terminal ends and internal interface with disulfide bonds, we found that the multimeric protein ADH cyclized into a robust monomeric-like form, resulting in superior thermostability compared to traditional protein engineering. This intersubunit cyclization approach is efficient and easy to perform, providing a novel method for engineering many important classes of multimeric proteins.
Topics: Actinobacteria; Alcohol Dehydrogenase; Bacterial Proteins; Cyclization; Enzyme Stability; Models, Molecular; Protein Conformation; Protein Multimerization; Temperature
PubMed: 28710270
DOI: 10.1128/AEM.01239-17 -
Environmental Health and Preventive... May 2016Many East Asians have the genetic polymorphisms rs1229984 in alcohol dehydrogenase 1B (ADH1B) and rs671 in aldehyde dehydrogenase 2 (ALDH2). Here we analyzed the...
Relationships of alcohol dehydrogenase 1B (ADH1B) and aldehyde dehydrogenase 2 (ALDH2) genotypes with alcohol sensitivity, drinking behavior and problem drinking in Japanese older men.
OBJECTIVES
Many East Asians have the genetic polymorphisms rs1229984 in alcohol dehydrogenase 1B (ADH1B) and rs671 in aldehyde dehydrogenase 2 (ALDH2). Here we analyzed the relationships of the two genotypes with alcohol sensitivity, drinking behavior and problem drinking among older and younger men living in rural areas of Japan.
METHODS
The subjects were 718 Japanese men aged 63.3 ± 10.8 (mean ± SD), categorized into the older (≥65 years, n = 357) and younger (<65 years, n = 361) groups. Facial flushing frequency, drinking behavior and positive CAGE results were compared among the genotypes using Bonferroni-corrected χ(2) test and a multivariate logistic regression analysis adjusting for age, BMI and lifestyle factors.
RESULTS
The frequency of 'always' facial flushing among the ADH1B*1/*2 carriers was significantly lower than that among the ADH1B*2/*2 carriers in the older group (P < 0.01). The alcohol consumption (unit/day) in the ADH1B*1/*2 carriers tended to be higher compared with that in the ADH1B*2/*2 carriers among the older group (P = 0.050). In the younger group, no significant differences in alcohol sensitivity and drinking habits were generally found among the ADH1B genotypes. The ADH1B*1/*1 genotype tended to be positively associated with problem drinking in the older group (P = 0.080) but not in the younger group. The ALDH2 genotypes consistently and strongly affected the alcohol sensitivity, drinking behavior and problem drinking in both the younger and older group.
CONCLUSIONS
We for the first time observed a significant difference in alcohol sensitivity between ADH1B*1/*2 and ADH1B*2/*2 in older men aged 65 and above.
Topics: Adult; Aged; Aged, 80 and over; Alcohol Dehydrogenase; Alcohol Drinking; Alcoholism; Aldehyde Dehydrogenase, Mitochondrial; Face; Genotype; Humans; Japan; Male; Middle Aged; Polymorphism, Genetic; Rural Population
PubMed: 26825972
DOI: 10.1007/s12199-016-0507-5 -
Nature Communications Oct 2019Aldehyde-alcohol dehydrogenase (AdhE) is a key enzyme in bacterial fermentation, converting acetyl-CoA to ethanol, via two consecutive catalytic reactions. Here, we...
Aldehyde-alcohol dehydrogenase (AdhE) is a key enzyme in bacterial fermentation, converting acetyl-CoA to ethanol, via two consecutive catalytic reactions. Here, we present a 3.5 Å resolution cryo-EM structure of full-length AdhE revealing a high-order spirosome architecture. The structure shows that the aldehyde dehydrogenase (ALDH) and alcohol dehydrogenase (ADH) active sites reside at the outer surface and the inner surface of the spirosome respectively, thus topologically separating these two activities. Furthermore, mutations disrupting the helical structure abrogate enzymatic activity, implying that formation of the spirosome structure is critical for AdhE activity. In addition, we show that this spirosome structure undergoes conformational change in the presence of cofactors. This work presents the atomic resolution structure of AdhE and suggests that the high-order helical structure regulates its enzymatic activity.
Topics: Acetyl Coenzyme A; Alcohol Dehydrogenase; Aldehyde Oxidoreductases; Cryoelectron Microscopy; Enzyme Assays; Escherichia coli Proteins; Ethanol; Mutation; Protein Conformation, alpha-Helical; Recombinant Proteins
PubMed: 31586059
DOI: 10.1038/s41467-019-12427-8 -
Proceedings of the National Academy of... Nov 1992Ethanol is the major metabolic product of glucose fermentation by the protozoan parasite Entamoeba histolytica under the anaerobic conditions found in the lumen of the... (Comparative Study)
Comparative Study
Ethanol is the major metabolic product of glucose fermentation by the protozoan parasite Entamoeba histolytica under the anaerobic conditions found in the lumen of the colon. Here an internal peptide sequence determined from a major 39-kDa amoeba protein isolated by isoelectric focusing followed by SDS/PAGE was used to clone the gene for the E. histolytica NADP(+)-dependent alcohol dehydrogenase (EhADH1; EC 1.1.1.2). The EhADH1 clone had an open reading frame that was 360 amino acids long and encoded a protein of approximately 39 kDa (calculated size). EhADH1 showed a 62% amino acid identity with the tetrameric NADP(+)-dependent alcohol dehydrogenase of Thermoanaerobium brockii. In contrast, EhADH1 showed a 15% amino acid identity with the closest human alcohol dehydrogenase. EhADH1 contained 18 of the 22 amino acids conserved in other alcohol dehydrogenases, including glycines involved in binding NAD(P)+ as well as histidine and cysteine residues involved in binding the catalytic zinc ion. Like the T. brockii alcohol dehydrogenase, EhADH1 lacked a 23-amino acid stretch present in other alcohol dehydrogenases that includes four cysteines that bind a second noncatalytic zinc ion. An EhADH1-glutathione-S-transferase fusion protein showed the expected NADP(+)-dependent alcohol dehydrogenase and NADPH-dependent acetaldehyde reductase activities. The enzymatic activities of the EhADH1 fusion protein were inhibited by pyrazole and 4-methylpyrazole.
Topics: Alcohol Dehydrogenase; Amino Acid Sequence; Animals; Base Sequence; Blotting, Southern; Cloning, Molecular; Electrophoresis, Gel, Two-Dimensional; Entamoeba histolytica; Genes; Glutathione Transferase; Kinetics; Molecular Sequence Data; NADP; Oligodeoxyribonucleotides; Phylogeny; Polymerase Chain Reaction; Recombinant Fusion Proteins; Sequence Homology, Amino Acid
PubMed: 1438208
DOI: 10.1073/pnas.89.21.10188