Enzymes containing zinc

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 linger proteins have an important regulatory function. In many of these systems, the non-redox-active Zn + ion is present as a Lewis 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. 

Carbonic anhydrase (CA), in particular, has proved a popular target for the development of structural models. The role of this enzyme is to catalyse the simple, but very important, reaction of CO2 fixation (Equation 12.1). 

A successful structural model for this process has been developed, which involves the tripodal tris(pyrazolyl)borate anion, resulting in the distorted tetrahedral species 12.20. An analogue of the key hydrogen carbonate catalytic intermediate in which the Zn + centre is five-coordinate is also known in the form of a bidentate nitrate analogue [Zn 77LHB(3-Phpz)3 (77 -N03)]. 

In mammals the bicarbonate ion, IICOT, is used as a buffer in blood and is catalysed by the zinc-containing enzyme carbonic anhydrase  

The uncatalysed reaction is slow(k= 9.5 x 10-2 Lmol 1s 1 at 25°C), however, in the presence of carbonic anhydrase the rate increases to 5 x 107 Lmol 1s 1 which represents 500,000 turnovers per second for each enzyme molecule. Carbonic anhydrase has a globular structure and has a mass of about 29 kDa. The single zinc ion is bound to three nitrogens (from histidine residues) and a water molecule or, as in Fig. 4.18, nearby amino acid occupies the fourth site. 

Experiments have shown that a metal is required for catalysis to occur but activity is seen for metals other than zinc. Its activity is inhibited by other small molecules that can bind to zinc in place of water and carbon dioxide, in particular cyanide, hydrogen sulfide and chloride that all bind tenaciously to transition metals. As well as its buffering ability, this enzyme represents a valuable method of converting carbon dioxide into carbonate so any advances in mimicking the behaviour of this enzyme may have implications for carbon storage. Conceivably carbon dioxide could be passed through a vat containing an aqueous solution of a carbonic 

As with carbonic anhydrase the metal is in a cleft that exposes the active site. Interestingly the metal-free enzyme is inactive but the cobalt and nickel analogues are more active. It appears that the transition state complex, where the terminal amino acid side chain is held in place while the peptide bond is hydrolysed, requires six-fold co-ordination. The activation energy required to change from the tetrahedral to octahedral geometries is higher for zinc than the other metals. 

The final exemplars of a tetrahedral zinc complex are the transcription factors known as zinc fingers. These are proteins about 30 amino acids in length that recognize regions of DNA to be transcribed into RNA sequences. Zinc binds in particular 

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Carboxypeptidases are zinc-containing enzymes that catalyze the hydrolysis of polypeptides at the C-terminal peptide bond. The bovine enzyme form A is a monomeric protein comprising 307 amino acid residues. The structure was determined in the laboratory of William Lipscomb, Harvard University, in 1970 and later refined to 1.5 A resolution. Biochemical and x-ray studies have shown that the zinc atom is essential for catalysis by binding to the carbonyl oxygen of the substrate. This binding weakens the C =0 bond by... 

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. 

Binding of bidentate sulfur ligands can also be of importance in the inhibition of zinc containing enzymes. An aminopeptidase which is essential for vancomycin antibiotic resistance in... 

Cerklewski and Forbes 1976). Also, excess zinc protects zinc-containing enzymes like ALAS, ferrochelatase, and ALAD. In vivo, aqueous solution containing zinc administered to rats significantly reduced the genotoxic effects induced by lead (Kowalska-Wochna et al. 1988). It was postulated that zinc s protective action may be related to its functioning in DNA and RNA polymerases and consequent enhancement of cell repair processes. 

The observed normal isotope effect of 1.9 provides further evidence supporting the role of Asp55 as the general base. Namely, a normal isotope effect of 1.9 is most consistent with general base catalysis by an amino acid side chain, as inverse isotope effects are commonly observed when a zinc-bound water molecule, or hydroxide, is the attacking nucleophile. For example, the zinc-containing enzymes AMP deaminase [111], thermolysin [112], stromelysin [113], and a desuccinylase [114] are each believed to utilize a zinc-bound water as the nucleophile, and all of these reactions are characterized by an inverse deuterium isotope effect. This inverse isotope effect is thought to result from a dominant... 

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). 

In our quest to find suitable N,N,0- igands as models for the active sites of the facial 2-His-l-carboxylate triad in iron and zinc containing enzymes, heteroscorpionate ligands like bis(3,5-dimethylpyrazol-... 

Excessive retention of sodium ions in the body leads to increased release of antidiuretic hormone and a resulting increase in body water. Many diuretics such as chlorothiazide (i) and clopamide (2) inhibit sodium and chloride ion resorption in the kidney tubules and promote potassium depletion, whereas amiloride (J) and triamterene ( 4) diminish the excretion of potassium while causing a loss of sodium ions6. Other diuretics such as acetazolamide (5) and dichlorphenamide (6) inhibit the ion-exchange reaction catalysed by the zinc-containing enzyme carbonic an-hydrase. 

Aminopeptidases that catalyze the hydrolysis of cysteinyl peptides are known. The membrane-bound aminopeptidases are glycoproteins, usually with molecular weights of about 100,000 daltons. They appear to be metalloproteins, one of the better known being a zinc-containing enzyme. Other enzymes, such as the leucine aminopeptidase, are cytosolic but, at least in this case, are also zinc-containing. The substrate specificity of these enzymes varies but most are relatively nonspecific. 

Optically active compounds having a free thiol functionality are interesting molecules because they can serve as inhibitors of zinc-containing enzymes [31]. 

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