alcohol dehydrogenase

alcohol dehydrogenase
Crystallographic structure of the homodimer of human ADH5.[1]
Identifiers
EC number	1.1.1.1
CAS number	9031-72-5
IntEnz	IntEnz view
BRENDA	BRENDA entry
ExPASy	NiceZyme view
KEGG	KEGG entry
MetaCyc	metabolic pathway
PRIAM	profile
PDB	structures
Gene Ontology	AmiGO / EGO
Search PMC articles PubMed articles

Alcohol dehydrogenases (ADH) (EC 1.1.1.1) are a group of seven dehydrogenase enzymes that occur in many organisms and facilitate the interconversion between alcohols and aldehydes or ketones with the reduction of NAD+ to NADH. In humans and many other animals, they serve to break down alcohols which could otherwise be toxic; in yeast and many bacteria, some alcohol dehydrogenases catalyze the opposite reaction as part of fermentation.

Discovery

Alcohol dehydrogenase was first discovered in the mid-1960s in Drosophila melanogaster.^{[dubious – discuss] [2]} Alcohol dehydrogenase is a dimer with a mass of 80 kDa.^[3]

Properties

The alcohol dehydrogenases comprise a group of several isozymes that facilitate the conversion of toxic alcohols to aldehydes. In mammals this is a redox (reduction/oxidation) reaction involving the coenzyme nicotinamide adenine dinucleotide (NAD⁺)

Alcohol dehydrogenase is responsible for catalyzing oxidation of primary and secondary alcohols to aldehydes and ketones, respectively, and also can affect the reverse reaction. It does not work well with primary alcohols.^{[dubious – discuss]} Instead, it works the best with secondary and cyclic alcohols.^[2]

Oxidation of alcohol

Mechanism of action in humans

Steps

1. Binding of the coenzyme NAD⁺;
2. Binding of the alcohol substrate by coordination to zinc;
3. Deprotonation of His-51;
4. Deprotonation of nicotinamide ribose;
5. Deprotonation of Ser-48;
6. Deprotonation of the alcohol;
7. Hydride transfer from the alkoxide ion to NAD⁺, leading to NADH and a zinc bound aldehyde or ketone;
8. Release of the product aldehyde;

The mechanism in yeast and bacteria is the reverse of this reaction. These steps are supported through kinetic studies.^[3]

Involved subunits

The substrate is coordinated to the zinc and this enzyme has two zinc atoms per subunit. One is the active site, which is involved in catalysis. In the active site, the ligands are Cys-46, Cys-174,His-67 and one water molecule. The other subunit is involved with structure. In this mechanism, the hydride from the alcohol goes to NAD⁺. Crystal structures indicate that the His-51 deprotonates the nicotinamide ribose, which deprotonates Ser-48. Finally, Ser-48 deprotonates the alcohol, making it an aldehyde.^[3] From a mechanistic perspective, if the enzyme adds hydride to the re face of NAD+, the resulting hydrogen is incorporated into the pro-R position. Enzymes that add hydride to the re face are deemed Class A dehydrogenases.

Active site

The active site of alcohol dehydrogenase

The active site consists of a zinc atom, His-67, Cys-174, Cys-46, Ser-48, His-51, Ile-269, Val-292, Ala-317, and Phe-319. The zinc coordinates the substrate(alcohol). The zinc is coordinated by Cys-146, Cys-174, and His-67. Phe-319, Ala-317, His-51, Ile-269 and Val-292 stabilize NAD⁺ by forming hydrogen bonds. His-51 and Ile-269 form hydrogen bonds with the alcohols on nicotinamide ribose. Phe-319, Ala-317 and Val-292 form hydrogen bonds with the amide on NAD⁺.^[3]

Types

Human

In humans, ADH exists in multiple forms as a dimer and is encoded by at least seven different genes. There are five classes (I-V) of alcohol dehydrogenase, but the hepatic form that is primarily used in humans is class 1. Class 1 consists of A,B, and C subunits that are encoded by the genes ADH1A, ADH1B, and ADH1C.^[4] The enzyme is contained in the lining of the stomach and in the liver. It catalyzes the oxidation of ethanol to acetaldehyde:

CH₃CH₂OH + NAD⁺ → CH₃CHO + NADH + H⁺

This allows the consumption of alcoholic beverages, but its evolutionary purpose is probably the breakdown of alcohols naturally contained in foods or produced by bacteria in the digestive tract. Others believe that its evolutionary purpose is involved in vitamin A metabolism, as alcohols are relatively 'empty' calories (about half that of a similar amount of a carbohydrate such as glucose), providing little net nutritional benefit.^{[citation needed]}

Alcohol dehydrogenase is also involved in the toxicity of other types of alcohol: for instance, it oxidizes methanol to produce formaldehyde and ethylene glycol to ultimately yield glycolic and oxalic acids. Humans have at least six slightly different alcohol dehydrogenases. Each is a dimer (i.e., consists of two polypeptides), with each dimer containing two zinc ions Zn²⁺. One of those ions is crucial for the operation of the enzyme: it is located at the catalytic site and holds the hydroxyl group of the alcohol in place.

Alcohol dehydrogenase activity varies between men and women, between young and old, and among populations from different areas of the world. For example, young women are unable to process alcohol at the same rate as young men because they do not express the alcohol dehydrogenase as highly. Though, the inverse is true amongst the middle-aged.^[5] The level of activity may not only be dependent on level of expression but due to allelic diversity among the population. These allelic differences have been linked to region of origin. For example, populations from Europe have been found to express an allele for the alcohol dehydrogenase gene that makes it much more active than those found in populations from Asia or the Americas.

This may be a correlating evolution with the rise of aldehyde dehydrogenase, which has been suggested as one of the more recognizable recent evolutionary changes in humans (along with lactose tolerance) - in order to make water safe in cities too dense to use springs, Europeans fermented alcoholic (and hence antiseptic) beverages, while Asians typically boiled their water (creating, among other things, tea). The Europeans' relatively greater alcohol consumption increased selection for those who didn't suffer from violent alcohol flush response in European populations.^{[citation needed]}

The human genes that encode class II, III, IV, and V alcohol dehydrogenases are ADH4, ADH5, ADH7, and ADH6, respectively.

Yeast and bacteria

Unlike humans, yeast and bacteria do not ferment glucose to lactate. Instead, they ferment it to ethanol and CO₂. The overall reaction can be seen below.

Glucose + 2ADP +2Pi → 2 ethanol + 2CO₂ + 2ATP +2H₂O^[6]

In yeast and many bacteria, alcohol dehydrogenase plays an important part in fermentation: pyruvate resulting from glycolysis is converted to acetaldehyde and carbon dioxide, and the acetaldehyde is then reduced to ethanol by an alcohol dehydrogenase called ADH1. The purpose of this latter step is the regeneration of NAD+, so that the energy-generating glycolysis can continue. Humans exploit this process to produce alcoholic beverages, by letting yeast ferment various fruits or grains. It is interesting to note that yeast can produce and consume their own alcohol.

The main alcohol dehydrogenase in yeast is larger than the human one, consisting of four rather than just two subunits. It also contains zinc at its catalytic site. Together with the zinc-containing alcohol dehydrogenases of animals and humans, these enzymes from yeasts and many bacteria form the family of "long-chain"-alcohol dehydrogenases.

Brewer's yeast also has another alcohol dehydrogenase, ADH2, which evolved out of a duplicate version of the chromosome containing the ADH1 gene. ADH2 is used by the yeast to convert ethanol back into acetaldehyde, and it is only expressed when sugar concentration is low. Having these two enzymes allows yeast to produce alcohol when sugar is plentiful (and this alcohol then kills off competing microbes), and then continue with the oxidation of the alcohol once the sugar, and competition, is gone.^[7]

Alcohol Dehydrogenase from Brewer's Yeast can be easily purified as follows: Using a 25 mM Pyrophosphate Buffer containing 5 mM Zinc Chloride, 5 mM EDTA, and 0.5 mg/ml BSA, disrupt yeast cells with a bead mill. Perform an ammonium sulfate precipitation at 40%, and discard the pellet. Perform a second ammonium sulfate precipitation at 70%, resuspend the resulting pellet in 1 ml of buffer. Run on a Sephadex G-100 column; ADH will elute in first few fractions. The sample can then be concentrated with a high concentration of ammonium sulfate as there should be few other proteins present.

Iron-containing

A third family of alcohol dehydrogenases, unrelated to the above two, are iron-containing ones. They occur in bacteria, and an (apparently inactive) form has also been found in yeast^{[citation needed]}. In comparison to enzymes the above families, these enzymes are oxygen-sensitive.

Other types

A further class of alcohol dehydrogenases belongs to quinoenzymes and requires quinoid cofactors (e. g., pyrroloquinoline quinone, PQQ) as enzyme-bound electron acceptors. A typical example for this type of enzyme is methanol dehydrogenase of methylotrophic bacteria.

Applications

In fuel cells, alcohol dehydrogenases can be used to catalyze the breakdown of fuel for an ethanol fuel cell. Scientists at Saint Louis University have used carbon-supported alcohol dehydrogenase with poly(methylene green) as an anode, with a nafion membrane, to achieve about 50 μA/cm².^[8]

In biotransformation, alcohol dehydrogenases are often used for the synthesis of enantiomerically pure stereoisomers of chiral alcohols. In contrast to the chemical process, the enzymes yield directly the desired enatiomer of the alcohol by reduction of the corresponding ketone.

Clinical significance

Alcoholism

There have been studies showing that ADH may have an influence on the dependence on ethanol metabolism in alcoholics. Researchers have tentatively detected a few genes to be associated with alcoholism. If the variants of these genes encode slower metabolizing forms of ADH2 and ADH3, there is increased risk of alcoholism. The studies have found that mutations of ADH2 and ADH3 are related to alcoholism in Asian populations. However, research continues in order to identify the genes and their influence on alcoholism.^[9]

Drug dependence

Drug dependence is another problem associated with ADH, which researchers think might be linked to alcoholism. One particular study suggests that drug dependence has seven ADH genes associated with it. These results may lead to treatments that target these specific genes. However, more research is necessary.^[10]

Picture gallery

Alcohol Dehydrogenase.   Horse LADH (Liver Alcohol Dehydrogenase)

See also

• Ethanol metabolism
• Aldehyde dehydrogenase
• ADH2
• Oxidoreductase
• Ethanol
• Blood alcohol content for rates of metabolism.

References

External links

Wikimedia Commons has media related to: Alcohol dehydrogenase

• PDBsum has links to three-dimensional structures of various alcohol dehydrogenases contained in the Protein Data Bank
• ExPASy contains links to the alcohol dehydrogenase sequences in Swiss-Prot, to a Medline literature search about the enzyme, and to entries in other databases.
• Radio Free Genome created a musical score from a sequence of alcohol dehydrogenase. MP3 audio version and an open source version of the software used to create it is available.