Alcohol dehydrogenase

Related Reading

Classification of the ADHs

ADH (E.C. 1.1.1.1) is an oxidoreductase that can catalyze the reversible oxidation of alcohols to aldehydes or ketones, with the reduction NAD + or NADP + at the same time. ADH is a large class of enzymes that can be further divided into at least three different enzyme superfamily: medium-chain (MDR) dehydrogenase/reductase, short-chain dehydrogenase/reductase, and iron-activated ADHs. The MDR superfamily consists of 350 amino acid residues (dimers or tetramers), and each subunit has a catalytic domain and a domain that binds to nucleotide NAD + or NADP +. Many enzymes of the MDR family have zinc in their active sites and have a sequence motif known as the zinc-containing ADH signature: GHEX₂GX₅ (G, A) X₂ (I, V, A, C, S). The genes encoding classic ADHs include ADH1, ADH2, ADH3, ADH4 and ADH5. Other genes include SFA1, BDH1, BDH2, SOR1 putative SOR2, the cinnamyl ADH, CDH1, XDH1, ADH6 and ADH7.  

Localization

The envelope structure of eukaryotic cells is organized into complex compartments in order to perform various biological functions. Knowing the position of proteins in the microenvironment of these cells is extremely important for understanding their functions and interactions. In the late 1960s, it was known that there were at least three NAD-dependent ADHs in baker's yeast. Two of these enzymes are located in the cytosol (Adh1p and Adh2p), and Adh3p is located in the mitochondrial matrix. When studying the mechanism of mitochondrial oxidation of cytoplasmic NADH, Overkamp (2000) and Bakker (2000) obtained sufficient evidence that ADH3 is not the sole mitochondrial ADH. The study of Huh (2003) determined that Adh5p is located in the cytoplasm, and in addition to Adh3p, Adh4p is also a mitochondrial enzyme. At present, the location of Adh6p and Adh7p is unclear, because most of the available information on the properties of these two enzymes comes from expression monitoring at the mRNA level or overexpression studies.

Structure Solution Overview

Saccharomyces cerevisiae alcohol dehydrogenase I (ADH1) is a constitutive enzyme that reduces acetaldehyde to ethanol during glucose fermentation. The structure of ADH1 was determined by X-ray crystallography at 2.4 Å resolution. The asymmetric unit contains four different subunits arranged as similar dimers, called AB and CD, which are similar to the horse liver ADH (Figure 1). Each subunit in the dimer has a typical Rossmann fold coenzyme binding domain, that is, a six-strand parallel β- pleated sheet, and there are two helices on each side of the sheet, and the coenzyme binds at the carboxyl terminal end. The extensive interaction of the two coenzyme binding domains produces an extended β-sheet in the dimer. Each subunit also has a catalytic domain, which contains a zinc atom to which the alcohol binds and a remote structural zinc ring in a distant loop. The A and C subunits have a "closed" conformation and contain NAD and TFE bonded to the catalytic zinc in a "classical" coordination, while the B and D subunits have an "open" conformation, which has an alternative coordination, and there is no binding coenzyme. A unit cell contains three biological molecules, each of which is two different tetramers with AB: AB and CD: CD subunits. 

Figure 1. Stereoviews of one asymmetric unit, an AB dimer, and of the biologic AB:AB tetramer in a back-to-back orientation (Raj, S.B.; et al. 2014)

Zinc Content and Coordination

Each subunit in the crystal structure of yeast ADH1 contains "catalytic" zinc and "structural" zinc, and its function is currently unclear. Although the biochemical experiments to determine the number of zinc atoms in yeast ADH1 produced different results, crystallography clearly showed that there is two zinc in each subunit, and these zincs exist in tetrahedral coordination. In the closed conformation, one subunit (A or C) in each asymmetric dimer binds NAD and TFE with the catalytic zinc in a "classical" manner with Cys-43, His-66, Cys-153 and the oxygen of TFE. In the open conformation, the other subunit (B or D) coordinates the catalytic zinc in an alternative manner, to Cys-43, His-66, Glu-67, and Cys-153 with the oxygen of TFE in the second sphere. The overlay of the catalytic domains of subunits A and B show that when zinc moves about 2.6 Å from Glu-67, the coordination of zinc is reversed, accompanied by a small amount of movement of zinc ligands. The structure of human ADH3 can illustrate an intermediate in the mechanism after coenzyme binding and before substrate binding. Obviously, alternative coordination is common, and the active site zinc coordination is flexible, so scientists have proposed a mechanism, that is, using alcohol or aldehyde instead of zinc-bound water.

Figure 2. Two different types of coordination of the catalytic zinc (Raj, S.B.; et al. 2014)

Properties

Alcohol dehydrogenase is a dimer with a mass of 80kDa, including a set of isoenzymes, which can convert ethanol into acetaldehyde. In mammals, this is a redox reaction involving the coenzyme nicotinamide adenine dinucleotide (NAD+). Alcohol dehydrogenase is responsible for catalyzing the oxidation of primary and secondary alcohols into aldehydes and ketones, and can also affect their reverse reactions. But for primary alcohols, this catalytic effect is not strong, while for secondary alcohols and cyclic alcohols, the catalytic effect is strong. The optimal pH value of alcohol dehydrogenase is 7.0-10.0, the enzyme activity reaches the maximum when the pH value is 8.0, and the enzyme activity is relatively stable when the pH value is 7.0; the optimal temperature of ADH is 37℃ and the temperature is 30-40 The enzyme activity is relatively stable at ℃, and the enzyme activity drops sharply after the temperature exceeds 45℃.

Application

• Disease diagnosis

ADH in the human body is mainly produced in the liver, so liver disease may be related to serum ADH activity. The use of spectrophotometry to determine the activity of ADH in serum and clinical exploration of this index is of great significance in the diagnosis of liver disease.

• Alcohol concentration

In daily social life, people are inevitably exposed to alcohol. Traffic accidents caused by drunk driving are not uncommon. In addition, excessive drinking can cause alcoholism. In view of the serious consequences of drinking accidents and alcoholism, the rapid determination of ethanol concentration in plasma has very important clinical value in preventing accidents and early diagnosis and treatment of acute alcoholism.

• Catalyst

In the chemical industry, the catalytic properties of ADH are used to produce many raw materials and intermediate reactants. In the process of converting carbon dioxide into methanol, ADH plays an enzyme catalytic role.

Clinical Significance

• Alcoholism

Studies have shown that alcohol dehydrogenase may lead to alcoholism-dependent alcoholism in patients. Researchers initially detected a few genes that may be related to alcoholism. If the ADH2 and ADH3 encoded by these gene variants enter the slow metabolic form, it may increase the risk of alcohol abuse. The study found that mutant ADH2 and ADH3 are related to alcohol abuse in Asian populations. However, whether this is really the case requires further research.

• Drug dependence

Drug dependence is another problem with alcohol dehydrogenase, which researchers believe may be related to alcoholism. A special study showed that drug dependence is related to 7 alcohol dehydrogenase related genes. These results may help targeted treatment of these specific genes. However, this needs more in-depth research.