Metallothionein
Metallothionein is a family of cysteine-rich, low molecular weight proteins. They are localized to the membrane of the Golgi apparatus. MTs have the capacity to bind both physiological and xenobiotic heavy metals through the thiol group of its cysteine residues, which represent nearly 30% of its constituent amino acid residues.
MT was discovered in 1957 by Vallee and Margoshe from purification of a Cd-binding protein from horse renal cortex. MT plays a role in the protection against metal toxicity and oxidative stress, and is involved in zinc and copper regulation. There are four main isoforms expressed in humans : MT1, MT2, MT3, and MT4. In the human body, large quantities are synthesised primarily in the liver and kidneys. Their production is dependent on availability of the dietary minerals such as zinc, copper, and selenium, as well as the amino acids histidine and cysteine.
Metallothioneins are rich in thiols, causing them to bind a number of trace metals. Metallothionein binds several Zn ions. One of few eukaryotic proteins distinguished as having a role in substantial metal detoxification. Zinc and Cadmium are tetrahedrally coordinated to cysteine residues, each metallothionein protein molecule may bind up to 7 atoms of Zn or Cd. The biosynthesis of metallothionein appeared to have increased by several-fold throughout oxidative stress to shield the cells against cytotoxicity and DNA damage. Metallothionein biosynthesis can also be induced by certain agents or conditions, for example, hormones, pharmaceuticals, alcohols, other substance treatments and many more. Metallothionein is a cytoplasmic protein, in an adult liver, it is localized mainly in the cytoplasm. In human fetus, metallothionein is localized in hepatocyte nuclei.
Structure and classification
MTs are present in a vast range of taxonomic groups, ranging from prokaryotes, protozoa, plants, yeast, invertebrates and vertebrates.The MTs from this diverse taxonomic range represent a high-heterogeneity sequence and do not show general homology; in spite of this, homology is found inside some taxonomic groups.
From their primary structure, MTs have been classified by different methods. The first one dates from 1987, when Fowler et al., established three classes of MTs: Class I, including the MTs which show homology with horse MT, Class II, including the rest of the MTs with no homology with horse MT, and Class III, which includes phytochelatins, Cys-rich enzymatically synthesised peptides.
The second classification was performed by Binz and Kagi in 2001, and takes into account taxonomic parameters and the patterns of distribution of Cys residues along the MT sequence.
It results in a classification of 15 families for proteinaceous MTs. Family 15 contains the plant MTs, which in 2002 have been further classified by Cobbet and Goldsbrough into 4 Types depending on the distribution of their Cys residues and a Cys-devoid regions characteristic of plant MTs.
A table including the principal aspects of the two latter classifications is included.
| Family | Name | Sequence pattern | Example |
| 1 | Vertebrate | K-x-C-C-x-C-C-P-x-C | Mus musculus MT1 MDPNCSCTTGGSCACAGSCKCKECKCTSCKKCCSCCPVGCAKCAQGCVCKGSSEKCRCCA |
| 2 | Molluscan | C-x-C-x-C-T-G-x-C-x-C-x-C-x-C-K | Mytilus edulis 10MTIV MPAPCNCIETNVCICDTGCSGEGCRCGDACKCSGADCKCSGCKVVCKCSGSCACEGGCTGPSTCKCAPGCSCK |
| 3 | Crustacean | P--P-C-C-x-C-x-C | Homarus americanus MTH MPGPCCKDKCECAEGGCKTGCKCTSCRCAPCEKCTSGCKCPSKDECAKTCSKPCKCCP |
| 4 | Echinoderms | P-D-x-K-C-V-C-C-x-C-x-C-x-C-C-x-C-C-x-C-C | Strongylocentrotus purpuratus SpMTA MPDVKCVCCKEGKECACFGQDCCKTGECCKDGTCCGICTNAACKCANGCKCGSGCSCTEGNCAC |
| 5 | Diptera | C-G-x-C-x-C-x-Q-x-C-x-C-xD-C-x-C | Drosophila melanogaster MTNB MVCKGCGTNCQCSAQKCGDNCACNKDCQCVCKNGPKDQCCSNK |
| 6 | Nematoda | K-C-C-x-C-C | Caenorhabditis elegans MT1 MACKCDCKNKQCKCGDKCECSGDKCCEKYCCEEASEKKCCPAGCKGDCKCANCHCAEQKQCGDKTHQHQGTAAAH |
| 7 | Ciliate | x-C-C-C-x ? | Tetrahymena thermophila MTT1 MDKVNSCCCGVNAKPCCTDPNSGCCCVSKTDNCCKSDTKECCTGTGEGCKCVNCKCCKPQANCCCGVNAKPCCFDPNSGCCCVSKTNNCCKSD TKECCTGTGEGCKCTSCQCCKPVQQGCCCGDKAKACCTDPNSGCCCSNKANKCCDATSKQECQTCQCCK |
| 8 | Fungal 1 | C-G-C-S-x-C-x-C-x-C-x-C-S-x-C | Neurospora crassa MT MGDCGCSGASSCNCGSGCSCSNCGSK |
| 9 | Fungal 2 | --- | Candida glabrata MT2 MANDCKCPNGCSCPNCANGGCQCGDKCECKKQSCHGCGEQCKCGSHGSSCHGSCGCGDKCECK |
| 10 | Fungal 3 | --- | Candida glabrata MT2 MPEQVNCQYDCHCSNCACENTCNCCAKPACACTNSASNECSCQTCKCQTCKC |
| 11 | Fungal 4 | C-X-K-C-x-C-x-C-K-C | Yarrowia lipolytica MT3 MEFTTAMLGASLISTTSTQSKHNLVNNCCCSSSTSESSMPASCACTKCGCKTCKC |
| 12 | Fungal 5 | --- | Saccharomyces cerevisiae CUP1 MFSELINFQNEGHECQCQCGSCKNNEQCQKSCSCPTGCNSDDKCPCGNKSEETKKSCCSGK |
| 13 | Fungal 6 | --- | Saccharomyces cerevisiae CRS5 TVKICDCEGECCKDSCHCGSTCLPSCSGGEKCKCDHSTGSPQCKSCGEKCKCETTCTCEKSKCNCEKC |
| 14 | Procaryota | K-C-A-C-x-C-L-C | Synechococcus sp SmtA MTTVTQMKCACPHCLCIVSLNDAIMVDGKPYCSEVCANGTCKENSGCGHAGCGCGSA |
| 15 | Plant | -x-C--C---x-C-x- | |
| 15.1 | Plant MTs Type 1 | C-X-C-X- C-X-C-X- C-X-C-X-spacer-C-X-C-X- C-X-C-X- C-X-C-X | Pisum sativum MT MSGCGCGSSCNCGDSCKCNKRSSGLSYSEMETTETVILGVGPAKIQFEGAEMSAASEDGGCKCGDNCTCDPCNCK |
| 15.2 | Plant MTs Type 2 | C-C-X-C-X-C-X- C-X-C-X- C-X-C-X-spacer- C-X-C-X- C-X-C-X- C-X-C-X | Lycopersicon esculentum MT MSCCGGNCGCGSSCKCGNGCGGCKMYPDMSYTESSTTTETLVLGVGPEKTSFGAMEMGESPVAENGCKCGSDCKCNPCTCSK |
| 15.3 | Plant MTs Type 3 | --- | Arabidopsis thaliana MT3 MSSNCGSCDCADKTQCVKKGTSYTFDIVETQESYKEAMIMDVGAEENNANCKCKCGSSCSCVNCTCCPN |
| 15.4 | Plant MTs Type 4 or Ec | C-x-C-X-C-X-C-X-C-X-C-X-HTTCGCGEHC- X-C-X-CSCGAXCNCASC-X | Triticum aestivum MT MGCNDKCGCAVPCPGGTGCRCTSARSDAAAGEHTTCGCGEHCGCNPCACGREGTPSGRANRRANCSCGAACNCASCGSTTA |
| 99 | Phytochelatins and other non-proteinaceous MT-like polypeptides | --- | Schizosaccharomyces pombe γEC-γEC-γECG |
More data on this classification are discoverable at the Expasy metallothionein page.
Secondary structure elements have been observed in several MTs SmtA from Syneccochoccus, mammalian MT3, Echinoderma SpMTA, fish Notothenia coriiceps MT, Crustacean MTH, but until this moment, the content of such structures is considered to be poor in MTs, and its functional influence is not considered.
Tertiary structure of MTs is also highly heterogeneous. While vertebrate, echinoderm and crustacean MTs show a bidominial structure with divalent metals as Zn or Cd, yeast and procariotyc MTs show a monodominial structure a bidominial structure similar to that of vertebrate MTs; 2) a codominial structure, in which two Cys-rich domains interact to form a single metallic cluster.
Quaternary structure has not been broadly considered for MTs. Dimerization and oligomerization processes have been observed and attributed to several molecular mechanisms, including intermolecular disulfide formation, bridging through metals bound by either Cys or His residues on different MTs, or inorganic phosphate-mediated interactions. Dimeric and polymeric MTs have been shown to acquire novel properties upon metal detoxification, but the physiological significance of these processes has been demonstrated only in the case of prokaryotic Synechococcus SmtA. The MT dimer produced by this organism forms structures similar to zinc fingers and has Zn-regulatory activity.
Metallothioneins have diverse metal-binding preferences, which have been associated with functional specificity. As an example, the mammalian Mus musculus MT1 preferentially binds divalent metal ions, Cd, while yeast CUP1 is selective for monovalent metal ions, Ag. Strictly metal-selective MTs with metal-specific physiological functions were discovered by Dallinger et al. in pulmonate snails. The Roman snail, for example, possesses a Cd-selective and a Cu-selective isoform involved in Cd detoxification and Cu regulation, respectively. While both isoforms contain unvaried numbers and positions of Cys residues responsible for metal ligation, metal selectivity is apparently achieved by sequence modulation of amino acid residues not directly involved in metal binding.
A novel functional classification of MTs as Zn- or Cu-thioneins is currently being developed based on these functional preferences.
Yeast
Metallothioneins are characterized by an abundance of cysteine residues and a lack of generic secondary structure motifs. Yeast Metallothionein are also alternatively named, Copper metallothionein. Yeast metallothionein coordinate much more strongly to Cu+ than Cu2+.Function
This protein functions in primary metal storage, transport, and detoxification. More specifically, Yeast MT stores copper so therefore protects the cell against copper toxicity by tightly chelating copper ions.For the first 40 residues in the protein the polypeptide wraps around the metal by forming two large parallel loops separated by a deep cleft containing the metal cluster.
Examples
Yeast MT can be found in the following:- Saccharomyces cerevisiae
- Neurospora crassa
Function
Metal binding
Metallothionein has been documented to bind a wide range of metals including cadmium, lead, zinc, mercury, copper, arsenic, silver, etc. Metalation of MT was previously reported to occur cooperatively but recent reports have provided strong evidence that metal-binding occurs via a sequential, noncooperative mechanism. The observation of partially metalated MT suggest that these species are biologically important.Metallothioneins likely participate in the uptake, transport, and regulation of zinc in biological systems. Mammalian MT binds three Zn ions in its beta domain and four in the alpha domain. Cysteine is a sulfur-containing amino acid, hence the name "-thionein". However, the participation of inorganic sulfide and chloride ions has been proposed for some MT forms. In some MTs, mostly bacterial, histidine participates in zinc binding. By binding and releasing zinc, metallothioneins may regulate zinc levels within the body. Zinc, in turn, is a key element for the activation and binding of certain transcription factors through its participation in the zinc finger region of the protein. Metallothionein also carries zinc ions from one part of the cell to another. When zinc enters a cell, it can be picked up by thionein and carried to another part of the cell where it is released to another organelle or protein. In this way thionein and metallothionein becomes a key component of the zinc signaling system in cells. This system is particularly important in the brain, where zinc signaling is prominent both between and within nerve cells. It also seems to be important for the regulation of the tumor suppressor protein p53.
Control of oxidative stress
Cysteine residues from MTs can capture harmful oxidant radicals like the superoxide and hydroxyl radicals. In this reaction, cysteine is oxidized to cystine, and the metal ions which were bound to cysteine are liberated to the media. As explained in the Expression and regulation section, this Zn can activate the synthesis of more MTs. This mechanism has been proposed to be an important mechanism in the control of the oxidative stress by MTs. The role of MTs in reducing oxidative stress has been confirmed by MT Knockout mutants, but some experiments propose also a prooxidant role for MTs.Metallothionein also plays a role in hematopoietic cell differentiation and proliferation, as well as prevention of apoptosis of early differentiated cells. Induced MT levels were adversely associated with sensitivity to etoposide-induced apoptosis, signifying that MT is a potential negative controller of apoptosis.
Expression and regulation
Metallothionein gene expression is induced by a high variety of stimuli, as metal exposure, oxidative stress, glucocorticoids, Vitamin D, hydric stress, fasting, exercise, etc. The level of the response to these inducers depends on the MT gene. MT genes present in their promoters specific sequences for the regulation of the expression, elements as metal response elements, glucocorticoid response elements, GC-rich boxes, basal level elements, and thyroid response elements.Metallothionein and disease
Cancer
Because MTs play an important role in transcription factor regulation, defects in MT function or expression may lead to malignant transformation of cells and ultimately cancer. Studies have found increased expression of MTs in some cancers of the breast, colon, kidney, liver, skin, lung, nasopharynx, ovary, prostate, mouth, salivary gland, testes, thyroid and urinary bladder; they have also found lower levels of MT expression in hepatocellular carcinoma and liver adenocarcinoma.There is evidence to suggest that higher levels of MT expression may also lead to resistance to chemotherapeutic drugs.
Autism
Heavy metal toxicity has been proposed as a hypothetical etiology of autism, and dysfunction of MT synthesis and activity may play a role in this. Many heavy metals, including mercury, lead, and arsenic have been linked to symptoms that resemble the neurological symptoms of autism. However, MT dysfunction has not specifically been linked to autistic spectrum disorders. A 2006 study, investigating children exposed to the vaccine preservative thiomersal, found that levels of MT and antibodies to MT in autistic children did not differ significantly from non-autistic children.A low zinc to copper ratio has been seen as a biomarker for autism and suggested as an indication that the Metallothionein system has been affected.
Further, there is indication that the mother's zinc levels may affect the developing baby's immunological state that may lead to autism and could be again an indication that the Metallothionein system has been affected.