Could function in retinol oxidation for the synthesis of retinoic acid, a hormone important for cellular differentiation. Medium-chain (octanol) and aromatic (m-nitrobenzaldehyde) compounds are the best substrates. Ethanol is not a good substrate but at the high ethanol concentrations reached in the digestive tract, it plays a role in the ethanol oxidation and contributes to the first pass ethanol metabolism.
A cDNA encoding a new type of alcohol dehydrogenase was cloned from a human stomach cDNA library. PCR amplification of 5'-stretch human stomach lambda gt11 library, using degenerate inosine-containing oligonucleotide probes compatible with peptide sequences of human sigma-ADH, resulted in a single product. Subsequently, internal non-degenerate primers were constructed according to the sequences occurring in the product. By PCR with combinations of these new primers and lambda gt11 forward and reverse primers, fragments of the cDNA containing its 5' and 3' ends were amplified. The full length cDNA sequence has 1125 nucleotides with a 72% similarity to those of human class I ADH. The polypeptide sequence, predicted from the cDNA, corresponds to 373 amino acids with a high degree of similarity (96%) to fragments of sigma-ADH previously reported. Northern hybridization analysis with the specific probe for the mRNA of this protein showed that it is expressed in the human stomach but not in the liver. These data indicate that the cDNA we cloned is that of human class IV ADH.
J. Biol. Chem. 270, 3625-3630 (1995)[PubMed:7876099]
A full-length 1966-base pair clone of the human class IV alcohol dehydrogenase (sigma-ADH) was isolated from a human stomach cDNA library. The 373-amino acid sigma-ADH encoded by this cDNA was expressed in Escherichia coli. The specific activity of the recombinant enzyme for ethanol oxidation at pH 7.5 and 25 degrees C, calculated from active-site titration of NADH binding, was 92 +/- 9 units/mg. Kinetic analysis of the catalytic efficiency (kcat/KM) of recombinant sigma-ADH for oxidation of primary alcohols indicated broad substrate specificity. Recombinant human sigma-ADH exhibited high catalytic efficiency for oxidation of all-trans-retinol to all-trans-retinal. This pathway is important in the synthesis of the transcriptional regulator all-trans-retinoic acid. Secondary alcohols and 3 beta-hydroxysteroids were inactive with sigma-ADH or were oxidized with very low efficiency. The KM of sigma-ADH for ethanol was 25 mM, and the KM for primary straight chain alcohols decreased substantially as chain length increased. There are important amino acid differences in the alcohol-binding site between the human class IV (sigma) and human class I (beta) alcohol dehydrogenases that appear to explain the high catalytic efficiency for all-trans-retinol, the high kcat for ethanol, and the low catalytic efficiency for secondary alcohols of sigma-ADH relative to beta 1-ADH. For example, modeling the binding of all-trans-retinol in the human beta 1-ADH structure suggested that coordination of retinol to the active-site zinc is hindered by a loop from residues 114 to 120 that is at the entrance to the alcohol-binding site. The deletion of Gly-117 in human sigma-ADH and a substitution of Leu for the bulky Tyr-110 appear to facilitate retinol access to the active-site zinc.
Kinetic constants of human class IV alcohol dehydrogenase (sigmasigma-ADH) support a role of the enzyme in retinoid metabolism, fatty acid omega-oxidation, and elimination of cytotoxic aldehydes produced by lipid peroxidation. Class IV is the human ADH form most efficient in the reduction of 4-hydroxynonenal (k(cat)/Km: 39,500 mM(-1) min(-1)). Class IV shows high activity with all-trans-retinol and 9-cis-retinol, while 13-cis-retinol is not a substrate but an inhibitor. Both all-trans-retinoic and 13-cis-retinoic acids are potent competitive inhibitors of retinol oxidation (Ki: 3-10 microM) which can be a basis for the regulation of the retinoic acid generation and of the pharmacological actions of the 13-cis-isomer. The inhibition of class IV retinol oxidation by ethanol (Ki: 6-10 mM) may be the origin of toxic and teratogenic effects of ethanol. H2-receptor antagonists are poor inhibitors of human and rat classes I and IV (Ki > 0.3 mM) suggesting a small interference in ethanol metabolism at the pharmacological doses of these common drugs.
J. Biol. Chem. 272, 18558-18563 (1997)[PubMed:9228021]
The structural determinants of substrate recognition in the human class IV, or sigmasigma, alcohol dehydrogenase (ADH) isoenzyme were examined through x-ray crystallography and site-directed mutagenesis. The crystal structure of sigmasigma ADH complexed with NAD+ and acetate was solved to 3-A resolution. The human beta1beta1 and sigmasigma ADH isoenzymes share 69% sequence identity and exhibit dramatically different kinetic properties. Differences in the amino acids at positions 57, 116, 141, 309, and 317 create a different topology within the sigmasigma substrate-binding pocket, relative to the beta1beta1 isoenzyme. The nicotinamide ring of the NAD(H) molecule, in the sigmasigma structure, appears to be twisted relative to its position in the beta1beta1 isoenzyme. In conjunction with movements of Thr-48 and Phe-93, this twist widens the substrate pocket in the vicinity of the catalytic zinc and may contribute to this isoenzyme's high Km for small substrates. The presence of Met-57, Met-141, and Phe-309 narrow the middle region of the sigmasigma substrate pocket and may explain the substantially decreased Km values with increased chain length of substrates in sigmasigma ADH. The kinetic properties of a mutant sigmasigma enzyme (sigma309L317A) suggest that widening the middle region of the substrate pocket increases Km by weakening the interactions between the enzyme and smaller substrates while not affecting the binding of longer alcohols, such as hexanol and retinol.
Interacting selectively and non-covalently with a nucleotide, any compound consisting of a nucleoside that is esterified with (ortho)phosphate or an oligophosphate at any hydroxyl group on the ribose or deoxyribose.
Kinetic constants of human class IV alcohol dehydrogenase (sigmasigma-ADH) support a role of the enzyme in retinoid metabolism, fatty acid omega-oxidation, and elimination of cytotoxic aldehydes produced by lipid peroxidation. Class IV is the human ADH form most efficient in the reduction of 4-hydroxynonenal (k(cat)/Km: 39,500 mM(-1) min(-1)). Class IV shows high activity with all-trans-retinol and 9-cis-retinol, while 13-cis-retinol is not a substrate but an inhibitor. Both all-trans-retinoic and 13-cis-retinoic acids are potent competitive inhibitors of retinol oxidation (Ki: 3-10 microM) which can be a basis for the regulation of the retinoic acid generation and of the pharmacological actions of the 13-cis-isomer. The inhibition of class IV retinol oxidation by ethanol (Ki: 6-10 mM) may be the origin of toxic and teratogenic effects of ethanol. H2-receptor antagonists are poor inhibitors of human and rat classes I and IV (Ki > 0.3 mM) suggesting a small interference in ethanol metabolism at the pharmacological doses of these common drugs.
Interacting selectively and non-covalently with retinol, vitamin A1, 2,6,6-trimethyl-1-(9'-hydroxy-3',7'-dimethylnona-1',3',5',7'-tetraenyl)cyclohex-1-ene, one of the three components that makes up vitamin A. Retinol is an intermediate in the vision cycle and it also plays a role in growth and differentiation.
J. Biol. Chem. 272, 18558-18563 (1997)[PubMed:9228021]
The structural determinants of substrate recognition in the human class IV, or sigmasigma, alcohol dehydrogenase (ADH) isoenzyme were examined through x-ray crystallography and site-directed mutagenesis. The crystal structure of sigmasigma ADH complexed with NAD+ and acetate was solved to 3-A resolution. The human beta1beta1 and sigmasigma ADH isoenzymes share 69% sequence identity and exhibit dramatically different kinetic properties. Differences in the amino acids at positions 57, 116, 141, 309, and 317 create a different topology within the sigmasigma substrate-binding pocket, relative to the beta1beta1 isoenzyme. The nicotinamide ring of the NAD(H) molecule, in the sigmasigma structure, appears to be twisted relative to its position in the beta1beta1 isoenzyme. In conjunction with movements of Thr-48 and Phe-93, this twist widens the substrate pocket in the vicinity of the catalytic zinc and may contribute to this isoenzyme's high Km for small substrates. The presence of Met-57, Met-141, and Phe-309 narrow the middle region of the sigmasigma substrate pocket and may explain the substantially decreased Km values with increased chain length of substrates in sigmasigma ADH. The kinetic properties of a mutant sigmasigma enzyme (sigma309L317A) suggest that widening the middle region of the substrate pocket increases Km by weakening the interactions between the enzyme and smaller substrates while not affecting the binding of longer alcohols, such as hexanol and retinol.
Molecular genetic studies have indicated that alcohol dehydrogenase may be involved in the synthesis of retinoic acid, a hormonal molecule regulating diverse cellular functions at the transcriptional level. Class IV alcohol dehydrogenase (ADH) has been reported to be the most efficient enzyme catalyzing oxidation of retinol in human ADH family. Initial velocity, product inhibition, and dead-end inhibition experiments were performed with the recombinant human class IV ADH to elucidate kinetic mechanism with all-trans-retinol and all-trans-retinal as natural substrates. Fluorescence quenching was titrated in formation of the binary and abortive ternary enzyme complexes. The minimal mechanism deduced from steady-state kinetic and equilibrium binding studies is best described as an asymmetric rapid equilibrium random mechanism with two dead-end ternary complexes for retinol oxidation and a rapid equilibrium ordered mechanism with one dead-end ternary complex for retinal reduction, a unique mechanistic form for zinc-containing ADHs in the medium chain dehydrogenase/reductase superfamily. Dissociation constants for the binary complexes as well as the productive and abortive ternary complexes determined from different experimental approaches are in reasonable agreement. Kinetic isotope effect studies suggest rate-limiting isomerization of the central ternary complexes in both reaction directions. The potential interference of retinol metabolism by ethanol through the ADH pathway may play a significant role in the pathogenesis of fetal alcohol syndrome and alcohol-related upper digestive tract cancer.
J. Biol. Chem. 270, 3625-3630 (1995)[PubMed:7876099]
A full-length 1966-base pair clone of the human class IV alcohol dehydrogenase (sigma-ADH) was isolated from a human stomach cDNA library. The 373-amino acid sigma-ADH encoded by this cDNA was expressed in Escherichia coli. The specific activity of the recombinant enzyme for ethanol oxidation at pH 7.5 and 25 degrees C, calculated from active-site titration of NADH binding, was 92 +/- 9 units/mg. Kinetic analysis of the catalytic efficiency (kcat/KM) of recombinant sigma-ADH for oxidation of primary alcohols indicated broad substrate specificity. Recombinant human sigma-ADH exhibited high catalytic efficiency for oxidation of all-trans-retinol to all-trans-retinal. This pathway is important in the synthesis of the transcriptional regulator all-trans-retinoic acid. Secondary alcohols and 3 beta-hydroxysteroids were inactive with sigma-ADH or were oxidized with very low efficiency. The KM of sigma-ADH for ethanol was 25 mM, and the KM for primary straight chain alcohols decreased substantially as chain length increased. There are important amino acid differences in the alcohol-binding site between the human class IV (sigma) and human class I (beta) alcohol dehydrogenases that appear to explain the high catalytic efficiency for all-trans-retinol, the high kcat for ethanol, and the low catalytic efficiency for secondary alcohols of sigma-ADH relative to beta 1-ADH. For example, modeling the binding of all-trans-retinol in the human beta 1-ADH structure suggested that coordination of retinol to the active-site zinc is hindered by a loop from residues 114 to 120 that is at the entrance to the alcohol-binding site. The deletion of Gly-117 in human sigma-ADH and a substitution of Leu for the bulky Tyr-110 appear to facilitate retinol access to the active-site zinc.
The chemical reactions and pathways resulting in the breakdown of ethanol, CH3-CH2-OH, a colorless, water-miscible, flammable liquid produced by alcoholic fermentation.
J. Biol. Chem. 270, 3625-3630 (1995)[PubMed:7876099]
A full-length 1966-base pair clone of the human class IV alcohol dehydrogenase (sigma-ADH) was isolated from a human stomach cDNA library. The 373-amino acid sigma-ADH encoded by this cDNA was expressed in Escherichia coli. The specific activity of the recombinant enzyme for ethanol oxidation at pH 7.5 and 25 degrees C, calculated from active-site titration of NADH binding, was 92 +/- 9 units/mg. Kinetic analysis of the catalytic efficiency (kcat/KM) of recombinant sigma-ADH for oxidation of primary alcohols indicated broad substrate specificity. Recombinant human sigma-ADH exhibited high catalytic efficiency for oxidation of all-trans-retinol to all-trans-retinal. This pathway is important in the synthesis of the transcriptional regulator all-trans-retinoic acid. Secondary alcohols and 3 beta-hydroxysteroids were inactive with sigma-ADH or were oxidized with very low efficiency. The KM of sigma-ADH for ethanol was 25 mM, and the KM for primary straight chain alcohols decreased substantially as chain length increased. There are important amino acid differences in the alcohol-binding site between the human class IV (sigma) and human class I (beta) alcohol dehydrogenases that appear to explain the high catalytic efficiency for all-trans-retinol, the high kcat for ethanol, and the low catalytic efficiency for secondary alcohols of sigma-ADH relative to beta 1-ADH. For example, modeling the binding of all-trans-retinol in the human beta 1-ADH structure suggested that coordination of retinol to the active-site zinc is hindered by a loop from residues 114 to 120 that is at the entrance to the alcohol-binding site. The deletion of Gly-117 in human sigma-ADH and a substitution of Leu for the bulky Tyr-110 appear to facilitate retinol access to the active-site zinc.
Sulfur mustard is a chemical warfare agent that causes blistering of the skin and damages the eyes and airway after environmental exposure. We have previously reported that thiodiglycol (TDG, 2,2'-bis-thiodiethanol), the hydrolysis product of sulfur mustard, is oxidized by alcohol dehydrogenase (ADH) purified from horse liver or present in mouse liver and human skin cytosol. Humans express four functional classes of ADH composed of several different isozymes, which vary in their tissue distribution, some occurring in skin. To help us evaluate the potential contribution of the various human isozymes toward toxicity in skin and in other tissues, we have compared the catalytic activity of purified human class I alphaalpha-, beta1beta1-, beta2beta2-, and gamma1gamma1-ADH, class II pi-ADH, class III chi-ADH, and class IV sigma-ADH with respect to TDG oxidation and their relative sensitivities to inhibition by pyrazole. Specific activities toward TDG were 123, 79, 347, 647, and 12 nmol/min/mg for the class I alphaalpha-, beta1,beta1-, beta2beta2-, and gamma1gamma1-ADH and class II pi-ADH, respectively. TDG was not a substrate for class III chi-ADH. The specific activity of class IV sigma-ADH was estimated at about 1630 nmol/min/mg. 1 mM pyrazole, a potent inhibitor of class I ADH, inhibited the class I alphaalpha, beta1beta1, beta2beta2, and gamma1gamma1 ADH and class IV sigma-ADH by 83, 100, 56, 90, and 73%, respectively. The class I alphaalpha- and beta1beta1-ADH oxidized TDG with kcat/Km value of 7-8 mM(-1) min(-1), beta2beta2-ADH with a value 19 mM(-1) min(-1) and class I gamma1gamma1-ADH with a value of 176 mM(-1) min(-1). The kcat/Km value for class IV sigma-ADH was estimated at 4 mM(-1) min(-1). The activities of class IV sigma-ADH and class I gamma1gamma1-ADH are of significant interest because of their prevalence in eyes, lungs, stomach, and skin, all target organs of sulfur mustard toxicity.
Kinetic constants of human class IV alcohol dehydrogenase (sigmasigma-ADH) support a role of the enzyme in retinoid metabolism, fatty acid omega-oxidation, and elimination of cytotoxic aldehydes produced by lipid peroxidation. Class IV is the human ADH form most efficient in the reduction of 4-hydroxynonenal (k(cat)/Km: 39,500 mM(-1) min(-1)). Class IV shows high activity with all-trans-retinol and 9-cis-retinol, while 13-cis-retinol is not a substrate but an inhibitor. Both all-trans-retinoic and 13-cis-retinoic acids are potent competitive inhibitors of retinol oxidation (Ki: 3-10 microM) which can be a basis for the regulation of the retinoic acid generation and of the pharmacological actions of the 13-cis-isomer. The inhibition of class IV retinol oxidation by ethanol (Ki: 6-10 mM) may be the origin of toxic and teratogenic effects of ethanol. H2-receptor antagonists are poor inhibitors of human and rat classes I and IV (Ki > 0.3 mM) suggesting a small interference in ethanol metabolism at the pharmacological doses of these common drugs.
A fatty acid oxidation process in which the methyl group at the end of the fatty acid molecule (the omega carbon) is first oxidized to a hydroxyl group, then to an oxo group, and finally to a carboxyl group. The long chain dicarboxylates derived from omega-oxidation then enter the beta-oxidation pathway for further degradation.
Kinetic constants of human class IV alcohol dehydrogenase (sigmasigma-ADH) support a role of the enzyme in retinoid metabolism, fatty acid omega-oxidation, and elimination of cytotoxic aldehydes produced by lipid peroxidation. Class IV is the human ADH form most efficient in the reduction of 4-hydroxynonenal (k(cat)/Km: 39,500 mM(-1) min(-1)). Class IV shows high activity with all-trans-retinol and 9-cis-retinol, while 13-cis-retinol is not a substrate but an inhibitor. Both all-trans-retinoic and 13-cis-retinoic acids are potent competitive inhibitors of retinol oxidation (Ki: 3-10 microM) which can be a basis for the regulation of the retinoic acid generation and of the pharmacological actions of the 13-cis-isomer. The inhibition of class IV retinol oxidation by ethanol (Ki: 6-10 mM) may be the origin of toxic and teratogenic effects of ethanol. H2-receptor antagonists are poor inhibitors of human and rat classes I and IV (Ki > 0.3 mM) suggesting a small interference in ethanol metabolism at the pharmacological doses of these common drugs.
A metabolic process that results in the removal or addition of one or more electrons to or from a substance, with or without the concomitant removal or addition of a proton or protons.
J. Biol. Chem. 270, 3625-3630 (1995)[PubMed:7876099]
A full-length 1966-base pair clone of the human class IV alcohol dehydrogenase (sigma-ADH) was isolated from a human stomach cDNA library. The 373-amino acid sigma-ADH encoded by this cDNA was expressed in Escherichia coli. The specific activity of the recombinant enzyme for ethanol oxidation at pH 7.5 and 25 degrees C, calculated from active-site titration of NADH binding, was 92 +/- 9 units/mg. Kinetic analysis of the catalytic efficiency (kcat/KM) of recombinant sigma-ADH for oxidation of primary alcohols indicated broad substrate specificity. Recombinant human sigma-ADH exhibited high catalytic efficiency for oxidation of all-trans-retinol to all-trans-retinal. This pathway is important in the synthesis of the transcriptional regulator all-trans-retinoic acid. Secondary alcohols and 3 beta-hydroxysteroids were inactive with sigma-ADH or were oxidized with very low efficiency. The KM of sigma-ADH for ethanol was 25 mM, and the KM for primary straight chain alcohols decreased substantially as chain length increased. There are important amino acid differences in the alcohol-binding site between the human class IV (sigma) and human class I (beta) alcohol dehydrogenases that appear to explain the high catalytic efficiency for all-trans-retinol, the high kcat for ethanol, and the low catalytic efficiency for secondary alcohols of sigma-ADH relative to beta 1-ADH. For example, modeling the binding of all-trans-retinol in the human beta 1-ADH structure suggested that coordination of retinol to the active-site zinc is hindered by a loop from residues 114 to 120 that is at the entrance to the alcohol-binding site. The deletion of Gly-117 in human sigma-ADH and a substitution of Leu for the bulky Tyr-110 appear to facilitate retinol access to the active-site zinc.
Any process that results in a change in state or activity of a cell or an organism (in terms of movement, secretion, enzyme production, gene expression, etc.) as a result of a stimulus from a bacterium.
Human gastric mucosa contains three alcohol dehydrogenase (ADH) isozymes (classes I, III, and IV). Various factors such as Helicobacter pylori infection, sex, age, and the part of the stomach involved have been suggested to affect alcohol dehydrogenase activities, although these views are controversial. In this study, these unsettled issues were reexamined.
Any process that results in a change in state or activity of a cell or an organism (in terms of movement, secretion, enzyme production, gene expression, etc.) as a result of an ethanol stimulus.
Kinetic constants of human class IV alcohol dehydrogenase (sigmasigma-ADH) support a role of the enzyme in retinoid metabolism, fatty acid omega-oxidation, and elimination of cytotoxic aldehydes produced by lipid peroxidation. Class IV is the human ADH form most efficient in the reduction of 4-hydroxynonenal (k(cat)/Km: 39,500 mM(-1) min(-1)). Class IV shows high activity with all-trans-retinol and 9-cis-retinol, while 13-cis-retinol is not a substrate but an inhibitor. Both all-trans-retinoic and 13-cis-retinoic acids are potent competitive inhibitors of retinol oxidation (Ki: 3-10 microM) which can be a basis for the regulation of the retinoic acid generation and of the pharmacological actions of the 13-cis-isomer. The inhibition of class IV retinol oxidation by ethanol (Ki: 6-10 mM) may be the origin of toxic and teratogenic effects of ethanol. H2-receptor antagonists are poor inhibitors of human and rat classes I and IV (Ki > 0.3 mM) suggesting a small interference in ethanol metabolism at the pharmacological doses of these common drugs.
The chemical reactions and pathways involving retinoids, any member of a class of isoprenoids that contain or are derived from four prenyl groups linked head-to-tail. Retinoids include retinol and retinal and structurally similar natural derivatives or synthetic compounds, but need not have vitamin A activity.
Kinetic constants of human class IV alcohol dehydrogenase (sigmasigma-ADH) support a role of the enzyme in retinoid metabolism, fatty acid omega-oxidation, and elimination of cytotoxic aldehydes produced by lipid peroxidation. Class IV is the human ADH form most efficient in the reduction of 4-hydroxynonenal (k(cat)/Km: 39,500 mM(-1) min(-1)). Class IV shows high activity with all-trans-retinol and 9-cis-retinol, while 13-cis-retinol is not a substrate but an inhibitor. Both all-trans-retinoic and 13-cis-retinoic acids are potent competitive inhibitors of retinol oxidation (Ki: 3-10 microM) which can be a basis for the regulation of the retinoic acid generation and of the pharmacological actions of the 13-cis-isomer. The inhibition of class IV retinol oxidation by ethanol (Ki: 6-10 mM) may be the origin of toxic and teratogenic effects of ethanol. H2-receptor antagonists are poor inhibitors of human and rat classes I and IV (Ki > 0.3 mM) suggesting a small interference in ethanol metabolism at the pharmacological doses of these common drugs.
There are 7 different ADH's isozymes in human: three belongs to class-I: alpha, beta, and gamma, one to class-II: pi, one to class-III: chi, one to class-IV: ADH7 and one to class-V: ADH6.
A reference proteome is a set of protein sequences derived from a complete proteome which constitutes a defined standard for a particular user community. Reference proteomes are manually defined according to a number of criteria. They cover the proteomes of well- studied model organisms and other proteomes of interest for biomedical and biotechnological research. Reference proteomes have been selected to provide broad coverage of the tree of life, and constitute a representative cross-section of the taxonomic diversity to be found within UniProtKB.
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