Iron

 Iron is one of the most functionally versatile of the physiologically essential transition metals. Both in hemoglobin and myoglobin the heme-bound Fe2+ iron is used to bind a diatomic gas, O2 , for transport and storage, respectively. Similarly, in marine invertebrates, the iron present in the diiron center of hemerythrin (Figure 1) can bind and transport oxygen. By contrast, the iron atoms contained in the heme groups of the b- and c-type cytochromes and the Fe-S clusters and Rieske iron centers (Figure 2) of other electron transport chain components transport electrons by cycling between their ferrous (+2) and ferric state (+3).

Fig1. Diiron center of the deoxy (left) and oxy (right) forms of hemerythrin. Shown are the side chains of the histidine, glutamate, and aspartate residues responsible for binding the metal ions to the polypeptide.

Fig2. Structure of a Rieske iron center. Rieske iron centers are a type of 2Fe-2S cluster in which histidine residues replace two of the cysteine residues that normally bind the prosthetic group to the polypeptide chain.

Roles of Iron in Redox Reactions

The iron atoms of many metalloproteins facilitate the catalysis of oxidation–reduction, or redox, reactions. Stearoyl-acyl carrier protein D9-desaturase and type 1 ribonucleotide reductase employ hemerythrin-like diiron centers to catalyze the reduction of carbon–carbon double bonds and an alcohol, respectively, to methylene groups. Methane monooxygenase uses a similar diiron center for the oxidation of methane to methanol. The members of the cytochrome P450 family generate Fe = O3+. This is a powerful oxidant that participates in the reduction and neutralization of a broad range of xenobiotics via the two-electron reduction of O2 , a complex process during which the heme iron cycles between several, that is, +2, +3, +4, and +5, oxidation states.

Participation of Iron in Non-Redox Reactions

 Purple acid phosphatases, bimetallic enzymes containing one atom of iron matched with a second metal, such as Zn, Mn, Mg, or another Fe, catalyze the hydrolysis of phosphomonoesters. Myeloperoxidase employs heme iron to catalyze the condensation of H₂O₂ with Cl⁻ ions to generate hypochlorous acid, HOCl, a potent bacteriocide used by macrophages to kill entrapped microorganisms. It recently has been shown that many enzymes involved in DNA replication and repair, including DNA helicase, DNA primase, several DNA polymerases, some glycosylases and endonucleases, and several transcription factors contain Fe-S clusters. While their elimination generally results in a loss of protein function, the specific role(s) performed by these Fe-S centers remains cryptic. However, since most are located in the DNA binding, rather than the catalytic, domains of these proteins, it has been proposed that these Fe-S centers may function as electrochemical detectors for the identification of damaged DNA. Others speculate that these clusters serve as redox-sensitive modulators of catalytic activity or DNA binding, or simply as stabilizers of the three-dimensional structure of these proteins.

Manganese

Humans contain a handful of Mn-containing enzymes, the majority of which are located within the mitochondria. These include isocitrate dehydrogenase from the tricarboxylic acid (TCA) cycle, two key players in nitrogen metabolism: glutamate synthetase and arginase, and the gluconeogenic enzymes pyruvate carboxylase and phosphoenolpyruvate carboxykinase, isopropyl malate synthase, and the mitochondrial iso zyme of superoxide dismutase. In most of these enzymes, Mn is present in the +2 oxidation state and is presumed to act as a Lewis acid. By contrast, some bacterial organisms employ Mn in several enzymes responsible for catalyzing redox reactions, where it cycles between the +2 and +3 oxidation states in, for example, Mn-superoxide dismutase (Mn-SOD), Mn-ribonucleotide reductase, and Mn-catalase.

Zinc

 Unlike the divalent (+2) ions of other first-row transition metals , the valence shell of Zn²⁺ possesses a full set of electrons. As a consequence, Zn²⁺ ions do not adopt alternate oxidation states under physiologic conditions, rendering it unsuitable to participate in electron transport processes or as a catalyst for redox reactions. On the other hand, redox inert Zn²⁺ ions also pose a minimal risk of generating harmful ROS species. Its unique status among the physiologically essential transition metals renders Zn²⁺ an ideal candidate as a ligand for stabilizing the protein conformation.

It has been estimated that the human body contains 3000 zinc-containing metalloproteins. The vast majority of these are transcription factors and other DNA- and RNA-binding proteins that contain anywhere from one to thirty copies of a Zn²⁺ -containing polynucleotide-binding domain known as the zinc finger. Zinc fingers consist of a polypeptide loop whose conformation is stabilized by the interactions between Zn²⁺ and lone pairs of electrons donated by the sulfur and nitrogen atoms contained in two conserved cysteine and two conserved histidine residues. Zinc fingers bind polynucleotides with a high degree of site specificity that is conferred, at least in part, by variations in the sequence of amino acids that make up the remainder of the loop. Scientists are working to exploit this combination of small size and binding specificity to construct sequence-specific nucleases for use in genetic engineering and, eventually, gene therapy.

Zn²⁺ is also an essential component of several metalloen zymes, including carboxypeptidase A, carbonic anhydrase II, adenosine deaminase, alkaline phosphatase, phospholipase C, leucine aminopeptidase, the cytosolic form of superoxide dismutase, and alcohol dehydrogenase. Zn²⁺ is also a component of the type II β-lactamases used by bacteria to neutralize penicillin and other lactam antibiotics. These metalloenzymes exploit the Lewis acid properties of Zn²⁺ to stabilize the development of negatively charged intermediates, polarize the distribution of electrons in carbonyl groups, and enhance the nucleophilicity of water (Figure 3).

Fig3. Role of Zn²⁺in catalytic mechanisms of β-lactamase II. Zn²⁺ is bound to the enzyme via the nitrogen atoms present in the side chains of multiple histidine (H) residues. Left: Zn²⁺ activates a water molecule, one of whose protons is accommodated by aspartic acid (D) residue 120, which (middle) executes a nucleophilic attack on carbonyl C of the lactam ring of the antibiotic. Right: D120 then donates the bound proton to the lactam nitrogen, facilitating the cleavage of the C-N bond in the tetrahedral intermediate.

 Cobalt

The predominant and, thus far, only known biochemical function of dietary cobalt is as the core component of 5′-deoxy adenosylcobalamin, otherwise known as vitamin B12 . The Co3+ in this cofactor resides at the center of a tetrapyrrole corrin ring where it acts as a Lewis base that binds to and facilitates the transfer of one-carbon methyl or methylene groups. In humans, this includes the enzyme catalyzed transfer of a –CH3 group from tetrahydrofolate to homocysteine, the final step in the synthesis of the amino acid methionine , and the rearrangement of methylmalonyl-CoA to form succinyl-CoA during the catabolism of the propionate generated from the metabolism of isoleucine and lipids containing odd numbers of amino acids (see Figure 19–2). During the latter reaction, the Co3+ is transiently reduced to the 2+ oxidation state by abstracting an electron to generate a reactive methylene radical, R-CH2 •. More information of Co and vitamin B12 can be found in Chapter 44.

Copper

Copper is a functionally essential component of approximately  30 different metalloenzymes in humans, including cytochrome oxidase, dopamine β-hydroxylase, tyrosinase, the cytosolic form of superoxide dismutase (Cu, Zn-SOD), and lysyl oxidase. Dopamine β-hydroxylase and tyrosinase are both catecholamine oxidases, enzymes that oxidize the ortho position in the phenol rings of L-dopamine and tyrosine, respectively. The former is the final step in the pathway by which epinephrine is synthesized in the adrenal gland, while the latter is the first and rate-limiting step in the synthesis of melanin. Both dopamine β-hydroxylase and tyrosinase are members of the type-3 family of copper proteins, which share a common dicopper center. As shown in Figure 4, the copper atoms in catecholamine oxidases chelate a molecule of molecular oxygen, activating it for attack on a phenol ring. During this process the copper atoms cycle between the +2 and +1 oxidation states. Another type-3 copper protein is hemocyanin. Unlike the catecholamine oxidases, the dicopper center of hemocyanin serves to transport oxygen in invertebrate animals such as mollusks that lack hemoglobin.

In Cu, Zn-SOD, the Cu2+ atom in the bimetallic center abstracts an electron from superoxide, O2 −, an extremely reactive and cytotoxic ROS, forming O2 and Cu1+. The Cu atom in the enzyme is then restored to its original, +2, valence state by donating an electron to a second molecule of super oxide, generating H₂O₂ . While hydrogen peroxide also is a ROS, it is considerably less reactive than O₂⁻, a radical anion. Moreover, it can be subsequently converted to water and O2 through the action of a second detoxifying enzyme, catalase .

Lysyl oxidase employs a single atom of Cu²⁺ to convert the epsilon amino groups on lysine side chains in collagen or elastin to aldehydes using molecular oxygen. The aldehyde groups on the side chain of the resulting amino acid, allysine (2-amino-6-oxo-hexanoic acid), then chemically react with the side chains of other allysine or lysine residues on adjacent polypeptides to generate the chemical crosslinks essential to the exceptional tensile strength of mature collagen and elastin fibers. Another essential feature of the enzyme is the presence of a modified amino acid, 2,4,5-trihydroxyphenylalanine qui none, in the active site. This modification is generated by the autocatalytic oxidation of the side chain of a conserved tyrosine residue by lysyl oxidase itself.

Fig4. Reaction mechanism of the catecholamine oxidases.

Nickel

 A variety of nickel-containing enzymes are present in bacterial organisms. Examples include Ni, Fe hydrogenase and methyl coenzyme M reductase, which catalyze redox reactions; ace tyl-CoA synthase, which catalyzes a transferase reaction; and superoxide dismutase, which catalyzes a disproportionation reaction. Ni is a key component of urease, an enzyme found in bacteria, fungi, and plants . However, the molecular basis of the dietary requirement for nickel in humans and other mammals has yet to be discovered.

Molybdenum

 Catalytic Roles of Molybdopterin

Molybdenum is a key component of the phylogenetically universal cofactor molybdopterin . In animals, molybdopterin serves as a catalytically essential prosthetic group for many enzymes, including xanthine oxidase, aldehyde oxidase, and sulfite oxidase. Xanthine oxidase, which also contains flavin, catalyzes the final two oxidative steps in the pathway by which uric acid is synthesized from purine nucleotides: the oxidation of hypoxanthine to xanthine and the oxidation of xanthine to uric acid. Catalysis of this two-stage process is facilitated by the ability of the bound Mo atom to cycle among the +4, +5, and +6 valence states. In addition to molybdopterin and flavin, aldehyde oxidase also contains an Fe-S cluster. Its complex suite of pros thetic groups enables the enzyme to oxidize a broad range of substrates, including many heterocyclic organic compounds. It has therefore been suggested that aldehyde oxidase participates, like the cytochrome P450 system, in the detoxification of xenobiotics.

Iron & Molybdenum Metalloenzymes

The Fe- and Mo-containing metalloenzyme sulfite oxidase is located in the mitochondria, where it catalyzes the oxidation of the sulfite (SO₃ ²⁻) generated by the catabolism of sulfur containing biomolecules to sulfate, SO4 2−. As for xanthine oxidase, the ability of the molybdenum ion to transition between the +6, +5, and +4 oxidation states is critical to providing a catalytic route by which the two electrons removed from the sulfite molecule can be sequentially transferred to two molecules of cytochrome c, each of which can carry only a single electron (Figure 5). Mutations in any one of three genes— MOCS1,MOCS2, or GPNH—whose protein products catalyze key steps in the synthesis of molybdopterin can lead to sulfite oxidase deficiency. Individuals who suffer from this autosomal inherited inborn error of metabolism are incapable of breaking down the sulfur-containing amino acids cysteine and methionine. The resulting accumulation of these amino acids and their derivatives in neonatal blood and tissues produces severe physical deformities and brain damage that leads to intractable seizures, severe mental retardation, and—in most cases—death during early childhood.

Fig5. Reaction mechanism of sulfite oxidase showing oxidation states of enzyme-bound iron and molybdenum atoms.

Vanadium

 Although nutritionally essential, the role of vanadium in living organisms remains cryptic. No vanadium-containing cofactor has been identified to date. Vanadium is found throughout the body in both its +4, for example, HVO4 2–, H2 VO4–, etc., and +5 oxidation states, for example, VO2+, HVO3+, etc. Various plasma proteins are known to bind oxides of vanadium, including albumin, immunoglobulin G, and transferrin. It has also recently been discovered that vanadium is a key component of the secreted “glue” with which marine mussels secure themselves to rocks, pilings, and ship’s bottoms. Although vanadate, a phosphate analog, is known to inhibit protein-tyrosine phosphatases and alkaline phosphatase in vitro, it is unclear whether these interactions are of physiologic significance.

Chromium

 The role of Cr in humans remains unknown. In the 1950s, a Cr3+-containing “glucose tolerance factor” was isolated from brewer’s yeast whose laboratory effects implicated this transition metal as a cofactor in the regulation of glucose metabolism. However, decades of research have failed to uncover either a Cr-containing biomolecule or a Cr-related genetic disease in animals. Nevertheless, many persons continue to ingest Cr-containing dietary supplements, such as Cr3+-picolinate, for their alleged weight-loss properties.