Metallothionein class

Most terrestrial invertebrates have limited access to water and feed on solid matter. As a consequence, they take up most of their nutrients by ingestion of foodstuffs that are also the vehicle for ingestion of contaminants. Many of the class a , metals that are taken up are found in membrane-bound granules in the cells of the hepatopancreas, although uncertainties remain as to the initiation of granule formation. Other metals, such as the class b metal cadmium, may be in the granule or may be bound to a metallothionein type protein. 

Metallothioneins are an ubiquitous class of proteins with which the important roles of metal storage (Cu, Zn) and detoxification (Cd, Pb, Hg) are associated EXAFS studies have been carried out on a variety of metallothioneins including Zttj, ZUjCuj, ZiigCuj, Cd, and to show that metal atoms are exclusively... 

Metallothioneins (MTs) are low molecular mass, cysteine-rich metalbinding proteins, which have been divided into three classes (see Kagi ... 

Grill, E., Winnacker, E.-L. Zenk, M.H. (1987). Phytochelatins, a class of heavy metal binding peptides from plants are functionally analogous to metallothioneins. Proceedings of the National Academy of Sciences (USA) 84, 439-43. 

As the focus of this review is on copper-dioxygen chemistry, we shall briefly summarize major aspects of the active site chemistry of those proteins involved in 02 processing. The active site structure and chemistry of hemocyanin (He, 02 carrier) and tyrosinase (Tyr, monooxygenase) will be emphasized, since the chemical studies described herein are most relevant to their function. The major classes of these proteins and their origins, primary functions, and leading references are provided in Table 1. Other classes of copper proteins not included here are blue electron carriers [13], copper-thiolate proteins (metallothioneines) [17], and NO reductases (e.g., nitrite [NIR] [18] or nitrous oxide [19]). 

Metallothioneins are evolutionarily conserved in that they contain a high cysteine content and lack of aromatic amino acids. However, few invertebrate MTs have been characterized, and these can exhibit wide variation in noncysteine amino acid residues. Initially, MTs were classified according to their structural characteristics. Class I MTs consist of polypeptides with highly conserved cysteine residue sequences and closely resemble the equine renal MT. Mammalian MTs consist of 61-68 amino acids residues and the sequence is highly conserved with respect to the position of the cysteine residues (e.g., cys-x-cys, cys-x-y-cys, and cys-cys sequences, where x and y are noncysteine, non-aromatic amino acids). Class II MTs have less conserved cysteine residues and are distantly related to mammalian MTs. Class III MTs are defined as atypical and consist of enzymatically synthesized peptides such as phy-tochelatins and cadystins. This former classification scheme has been replaced by a more complex system to include the increasing number of identified isoforms. 

Several classes of molecules are involved in metal homeostasis. Metallothioneins (MTs) are a family of cysteine-rich metal-binding proteins that in mammals appear to function in Zn homeostasis and protect against heavy metal toxicity and oxidative stress60. Glutathione (GSH) is a sulfhydryl-rich tripeptide that is generally involved in the protection of cells against toxicants and in the metabolism of xenobiotics71. MTs and GSH are found in fish, and their expression and functions have been conveniently studied with piscine cell lines. 

Cadmium, zinc, and copper bind to metallothioneins in different oxidation states and stoichiometries. In class I metallothioneins, 20 cysteine residues occur all of which participate in metal-ion binding. In zinc-metallothionein, seven Zn2+ ions are coordinated by three cysteine ligands each. A similar stoichiometry occurs in cadmium-metallothionein [54, 55], Additionally, basic amino acids (lysine and arginine) are in the close neighborhood of the cysteine residues and it had been suggested that they participated in the metal-binding as well [56,57]. Two cysteine residues occur in a Cys-x-Cys motive while the third cysteine residue is offset by some amino acids [58]. 

Similar oligonuclear centers may occur in class II metallothioneins as well [66]. In this class, 8 copper ions are coordinated by 12 cysteine ligands (Fig. 12). Two of these eight copper ions do not contribute to the characteristic CD-spec-trum and may easily be removed by copper chelators [67]. Consequently, the... 

In mammalians these metallothioneins normally possess chain lengths of 61 amino acids, of which 20 are invariant cysteines residues [58,316], These 20 residues participate in the binding of the up to twelve metal ions. Class II metallothioneins show little or no phylogenetic homology to the metallothioneins of class I, for instance, in sea urchins [317] and yeast [318]. Class III metallothio-... 

Fig. 36. Amino acid sequence of equine class I metallothioneins. From Kojima and Kagi 1979 [316] with permission...

The remarkable structural feature of metallothioneins is the sevenfold repetition of the sequence Cys-X-Cys in class I metallothioneins [58]. These two cy-... 

MT have been subdivided into three classes, namely I, II, and III (Fowler et al. 1987), while Binz and Kagi (1999) allocated these compounds to several families. Subsequently, Richards and Beattie (1995), Richards et al. (1996, 1997) and Szpunar (2000) showed capillary zone electrophoresis to be a rapid and reliable method for analysis of this protein. By using a diverse combination of techniques, the metal complexes of metallothionein in rat liver and kidney were first characterized, with two major MT-isoforms (MT-1 and MT-2) being detected in liver, and one MT isoform in kidney (Polec et al. 2002). The order of afHn-ity of metal ions to MTs is Cd > Zn, Cu, Ag, Hg > Bi > Pb, though this may change depending on the tissue involved. MTs are considered to be low molecular-weight pro-... 

A major component of the intracellular protein thiol pool is the class of inducible, cysteine-rich metallothionein (MT) proteins, important in heavy metal detoxification in eucaryotes (52). Studies of a human head and neck carcinoma cell line revealed no difference in nonprotein sulfhydryl content between parental cells and cells 30-fold resistant to cisplatin, but the latter had twofold greater levels of total protein sulfhydryl content (127). Several human and murine tumor cell lines resistant to cisplatin showed increased expression of one metallothionein, MT Ila, and increased levels of the protein (68). In addition, mouse cells transfected with the gene encoding metallothionein Ila showed a 10-fold increase in the level of MT accompanied by a 4.4-fold level of resistance to cw-DDP (68). By contrast, treatment of rats with cisplatin followed by chromatographic resolution of kidney metallothioneins showed that platinum did not elute with the MT fractions (84). Moreover, pretreatment of rats with Cd-" to induce MT production had no effect on the metabolism of platinum in kidney or liver (84). In vitro studies of the relative affinity of Cu-metallothionein for a variety of metal ions revealed very weak binding of divalent platinum... 

All eukaryotes and some prokaryotes such as cyanobacteria synthesize metal binding proteins— known collectively as metallothioneins—in response to high concentrations of several metals including Cd. These are cysteine-rich, low molecular mass polypetides whose thiol functional groups have high affinity for soft cations such as Cd " " [36], All plants, including phytoplankton, synthesize class... 

Kagi JHR (1991) Overview of metallothionein. Methods Enzymol 205 613-626 Kagi JHR (1993) Evolution, structure and chemical activity of class I metal-lothioneins an overview. In Suzuki KT, Imura N, Kimura K (eds) Metallothionein III. Birkhauser, Basel, pp 29-55 Klaassen CD (1978) Effect of metallothionein on the hepatic disposition of metals. Am J Physiol 234 E47-E53... 

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