Zinc-containing enzymes alcohol dehydrogenase

As you can judge from Table A, transition metal cations are frequently found in enzymes. The Zn2+ ion alone is known to be a component of at least 70 different enzymes. One of these, referred to as "alcohol dehydrogenase," is concentrated in the liver, where it acts to break down alcohols. Another zinc-containing enzyme is involved in the normal functioning of oil glands in the skin, which accounts for the use of Zn2+ compounds in the treatment of acne. 

The ALDs are a subset of the superfamily of medium-chain dehydrogenases/reductases (MDR). They are widely distributed, cytosolic, zinc-containing enzymes that utilize the pyridine nucleotide [NAD(P)+] as the catalytic cofactor to reversibly catalyze the oxidation of alcohols to aldehydes in a variety of substrates. Both endobiotic and xenobiotic alcohols can serve as substrates. Examples include (72) ethanol, retinol, other aliphatic alcohols, lipid peroxidation products, and hydroxysteroids (73). 

Alcohol dehydrogenase is a zinc-containing enzyme that catalyzes the reversible conversion of acetaldehyde and ethanol using NADH / NAD+. See below. 

Fig. 1 Active site structures of four zinc-containing enzymes (a) carboxypeptidase A from bovine pancreas, (b) (3-lactamase from Bacteroides fragilis, (c) human carbonic anhydrase. and (d) horse liver alcohol dehydrogenase.

There are two main routes for ethanol metabolism in the liver. The first and most important of these begins with the zinc-containing, qdoplasmic, alcohol dehydrogenase (ADH) group of enzymes (especially alcohol dehydrogenase IB). These enzymes are not particularly specific as they can also metabolise longer-chain alcohols produced in the co-oxidation pathway for fatty acids. ADH converts alcohol to acetaldehyde with a Km of 1.6 mmol/L ... 

Uncovering of the three dimentional structure of catalytic groups at the active site of an enzyme allows to theorize the catalytic mechanism, and the theory accelerates the designing of model systems. Examples of such enzymes are zinc ion containing carboxypeptidase A 1-5) and carbonic anhydrase6-11. There are many other zinc enzymes with a variety of catalytic functions. For example, alcohol dehydrogenase is also a zinc enzyme and the subject of intensive model studies. However, the topics of this review will be confined to the model studies of the former hydrolytic metallo-enzymes. 

One of the most important metals with regard to its role in enzyme chemistry is zinc. There are several significant enzymes that contain the metal, among which are carboxypeptidase A and B, alkaline phosphatase, alcohol dehydrogenase, aldolase, and carbonic anhydrase. Although most of these enzymes are involved in catalyzing biochemical reactions, carbonic anhydrase is involved in a process that is inorganic in nature. That reaction can be shown as... 

The inactivation of enzymes containing the zinc-thiolate moieties by peroxynitrite may initiate an important pathophysiological process. In 1995, Crow et al. [129] showed that peroxynitrite disrupts the zinc-thiolate center of yeast alcohol dehydrogenase with the rate constant of 3.9 + 1.3 x 1051 mol-1 s-1, yielding the zinc release and enzyme inactivation. Later on, it has been shown [130] that only one zinc atom from the two present in the alcohol dehydrogenase monomer is released in the reaction with peroxynitrite. Recently, Zou et al. [131] reported the same reaction of peroxynitrite with endothelial NO synthase, which is accompanied by the zinc release from the zinc-thiolate cluster and probably the formation of disulfide bonds between enzyme monomers. The destruction of zinc-thiolate cluster resulted in a decrease in NO synthesis and an increase in superoxide production. It has been proposed that such a process might be the mechanism of vascular disease development, which is enhanced by diabetes mellitus. 

Another zinc-utilizing enzyme is carbonate/dehydratase C (Kannan et al., 1972). Here, the zinc is firmly bound by three histidyl side chains and a water molecule or a hydroxyl ion (Fig. 27). The coordination is that of a distorted tetrahedron. Metals such as Cu(II), Co(Il), and Mn(ll) bind at the same site as zinc. Hg(II) also binds near, but not precisely at, this site (Kannan et al., 1972). Horse liver alcohol dehydrogenase (Schneider et al., 1983) contains two zinc sites, one catalytic and one noncatalytic. X-Ray studies showed that the catalytic Zn(II), bound tetrahedrally to two cysteines, one histidine, and water (or hydroxyl), can be replaced by Co(II) and that the tetrahedral geometry is maintained. This is also true with Ni(Il). Insulin also binds zinc (Adams etai, 1969 Bordas etal., 1983) and forms rhombohedral 2Zn insulin crystals. The coordination of the zinc consists of three symmetry-related histidines (from BIO) and three symmetry-related water molecules. These give an octahedral complex... 

Zinc is an essential trace element. More than 300 enzymes that require zinc ions for activity are known. Most catalyze hydrolysis reactions, but zinc-containing representatives of aU enzyme classes are known, such as, for instance, alcohol dehydrogenase (an oxidoreductase), famesyl-Zgeranyl transferase (a transferase), -lactamase (a hydrolase), carbonic anhydrase (a lyase) and phosphomannose isomerase. 

Although zinc itself is not redox-active, some class I enzymes containing zinc in their active sites are known. The most prominent are probably alcohol dehydrogenase and copper-zinc superoxide dismutase (Cu,Zn-SOD). AU have in common that the redox-active agent is another transition-metal ion (copper in Cu,Zn-SOD) or a cofactor such as nicotinamide adenine dinucleotide (NAD+/NADH). The Zn(II) ion affects the redox reaction only in an indirect manner, but is nevCTtheless essential and cannot be regarded simply as a structural factor. 

The latest proposed mechanisms1462 for several zinc-containing metalloenzymes combine elements from both types of mechanism by suggesting that the substrate binds to the enzyme through the C—O group, but that in the process the metal-bound water molecule is not displaced, so that the reaction proceeds via a five-coordinate intermediate. This hybrid mechanism is discussed below in greater detail for alcohol dehydrogenase. 

The NAD+-dependent alcohol dehydrogenase from horse liver contains one catalytically essential zinc ion at each of its two active sites. An essential feature of the enzymic catalysis appears to involve direct coordination of the enzyme-bound zinc by the carbonyl and hydroxyl groups of the aldehyde and alcohol substrates. Polarization of the carbonyl group by the metal ion should assist nucleophilic attack by hydride ion. A number of studies have confirmed this view. Zinc(II) catalyzes the reduction of l,10-phenanthroline-2-carbaldehyde by lV-propyl-l,4-dihy-dronicotinamide in acetonitrile,526 and provides an interesting model reaction for alcohol dehydrogenase (Scheme 45). The model reaction proceeds by direct hydrogen transfer and is absolutely dependent on the presence of zinc(II). The zinc(II) ion also catalyzes the reduction of 2- and 4-pyridinecarbaldehyde by Et4N BH4-.526 The zinc complex of the 2-aldehyde is reduced at least 7 x 105 times faster than the free aldehyde, whereas the zinc complex of the 4-aldehyde is reduced only 102 times faster than the free aldehyde. A direct interaction of zinc(II) with the carbonyl function is clearly required for marked catalytic effects to be observed. 

We have already seen a number of models for the zinc(II) containing enzymes such as carbonic anhydrase in Section 11.3.2. Zinc is an essential component in biochemistry, and forms part of the active site of more then 100 enzymes, of which hydrolases (such as alkaline phosphatase and carboxypeptidase A), transferases (e.g. DNA and RNA polymerase), oxidoreductases (e.g. alcohol dehydrogenase and superoxide dismutase) and lysases (carbonic anhydrase) are the most common. In addition, the non-enzyme zinc finger proteins have an important regulatory function. In many of these systems, the non-redox-active Zn2+ ion is present as a Fewis acidic centre at which substrates are coordinated, polarised and hence activated. Other roles of zinc include acting as a template and playing a structural or regulatory role. 

Substitution of foreign metals for the metals in metalloenzymes (those that contain metals as part of their structures) is an important mode of toxic action by metals. A common mechanism for cadmium toxicity is the substitution of this metal for zinc, a metal that is present in many metalloenzymes. This substitution occurs readily because of the chemical similarities between the two metals (for example, Cd2+ and Zn2+ behave alike in solution). Despite their chemical similarities, however, cadmium does not fulfill the biochemical function of zinc and a toxic effect results. Some enzymes that are affected adversely by the substitution of cadmium for zinc are adenosine triphosphate, alcohol dehydrogenase, and carbonic anhydrase. 

The major mechanistic difference between the pro-5 and the pro-/ specific enzymes in this area where thermodynamic constraints are weak or non-existent seems to be that the pro-/ specific enzymes contain a zinc ion at the active site whereas the pro-5 specific enzymes do not (Schneider-Bernlohr et al., 1986). In the mechanism of an NAD+-linked alcohol dehydrogenase shown in Scheme 6, in the reduction direction the substrate carbonyl group was shown as polarised by partial proton donation from a Bronsted acid BH + this polarisation can equally well be achieved by coordination to an active site zinc, which acts as a Lewis acid. One thus has two mechanistic classes of enzyme, but even this difference affects the stereochemistry only in a very limited region close to the break-point. 

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