Metallothionein

Metallothionein superfamily, eukaryotic
Solution structure of the beta-E-domain of wheat Ec-1 metallothionein.[1]
Identifiers
Symbol	Metallothionein_sfam
Pfam	PF00131
InterPro	IPR003019
Available protein structures: Pfam structures PDB RCSB PDB; PDBe; PDBj PDBsum structure summary

Yeast_MT
ag-substituted metallothionein from saccharomyces cerevisiae, nmr, minimized average structure
Identifiers
Symbol	Yeast_MT
Pfam	PF11403
InterPro	IPR022710
Available protein structures: Pfam structures PDB RCSB PDB; PDBe; PDBj PDBsum structure summary

Metallothionein (MT) is a family of cysteine-rich, low molecular weight (MW ranging from 500 to 14000 Da) proteins. They are localized to the membrane of the Golgi apparatus. MTs have the capacity to bind both physiological (such as zinc, copper, selenium) and xenobiotic (such as cadmium, mercury, silver, arsenic) heavy metals through the thiol group of its cysteine residues, which represent nearly 30% of its constituent amino acid residues.^[2]

MT was discovered in 1957 by Vallee and Margoshe from purification of a Cd-binding protein from horse (equine) renal cortex.^[3] MTs function is not clear, but experimental data suggest MTs may provide protection against metal toxicity, be involved in regulation of physiological metals (Zn and Cu) and provide protection against oxidative stress.^[4] There are four main isoforms expressed in humans (family 1, see chart below): MT1 (subtypes A, B, E, F, G, H, L, M, X), MT2, MT3, 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, as zinc, copper and selenium, and the amino acids histidine and cysteine.

Structure and classification

MTs are present in a vast range of taxonomic groups, ranging from prokaryotes (such as the cyanobacteria Syneccococus spp....), protozoa (p. ex. the ciliate Tetrahymena genera...), plants (such as Pisum sativum, Triticum durum, Zea mays, Quercus suber...), yeast (such as Saccharomyces cerevisiae, Candida albicans,...), invertebrates (such as the nematode Caenorhabditis elegans, the insect Drosophila melanogaster, the mollusc Mytilus edulis, or the echinoderm Strongylocentrotus purpuratus) and vertebrates (such as the chicken, Gallus gallus, or the mammalian Homo sapiens or Mus musculus).

The MTs from this diverse taxonomic range represent a high-heterogeneity sequence (regarding molecular weight and number and distribution of Cys residues) and do not show general homology; in spite of this, homology is found inside some taxonomic groups (such as vertebrate MTs).

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 (1, 2, 3 and 4) depending on the distribution of their Cys residues and a Cys-devoid regions (called spacers) 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(1,2)-C-C-x-C-C-P-x(2)-C	M.musculus MT1 MDPNCSCTTGGSCACAGSCKCKECKCTSCKKCCSCCPVGCAKCAQGCVCKGSSEKCRCCA
2	Molluscan	C-x-C-x(3)-C-T-G-x(3)-C-x-C-x(3)-C-x-C-K	M.edulis 10MTIV MPAPCNCIETNVCICDTGCSGEGCRCGDACKCSGADCKCSGCKVVCKCSGSCACEGGCTGPSTCKCAPGCSCK
3	Crustacean	P-[GD)-P-C-C-x(3,4)-C-x-C	H.americanus MTH MPGPCCKDKCECAEGGCKTGCKCTSCRCAPCEKCTSGCKCPSKDECAKTCSKPCKCCP
4	Echinoderms	P-D-x-K-C-[V,F)-C-C-x(5)-C-x-C-x(4)- C-C-x(4)-C-C-x(4,6)-C-C	S.purpuratus SpMTA MPDVKCVCCKEGKECACFGQDCCKTGECCKDGTCCGICTNAACKCANGCKCGSGCSCTEGNCAC
5	Diptera	C-G-x(2)-C-x-C-x(2)-Q-x(5)-C-x-C-x(2)D-C-x-C	D.melanogaster MTNB MVCKGCGTNCQCSAQKCGDNCACNKDCQCVCKNGPKDQCCSNK
6	Nematoda	K-C-C-x(3)-C-C	C.elegans MT1 MACKCDCKNKQCKCGDKCECSGDKCCEKYCCEEASEKKCCPAGCKGDCKCANCHCAEQKQCGDKTHQHQGTAAAH
7	Ciliate	x-C-C-C-x ?	T.termophila MTT1 MDKVNSCCCGVNAKPCCTDPNSGCCCVSKTDNCCKSDTKECCTGTGEGCKCVNCKCCKPQANCCCGVNAKPCCFDPNSGCCCVSKTNNCCKSD TKECCTGTGEGCKCTSCQCCKPVQQGCCCGDKAKACCTDPNSGCCCSNKANKCCDATSKQECQTCQCCK
8	Fungal 1	C-G-C-S-x(4)-C-x-C-x(3,4)-C-x-C-S-x-C	N.crassa MT MGDCGCSGASSCNCGSGCSCSNCGSK
9	Fungal 2	---	C.glabrata MT2 MANDCKCPNGCSCPNCANGGCQCGDKCECKKQSCHGCGEQCKCGSHGSSCHGSCGCGDKCECK
10	Fungal 3	---	C.glabrata MT2 MPEQVNCQYDCHCSNCACENTCNCCAKPACACTNSASNECSCQTCKCQTCKC
11	Fungal 4	C-X-K-C-x-C-x(2)-C-K-C	Y.lipolitica MT3 MEFTTAMLGASLISTTSTQSKHNLVNNCCCSSSTSESSMPASCACTKCGCKTCKC
12	Fungal 5	---	S.cerevisiae CUP1 MFSELINFQNEGHECQCQCGSCKNNEQCQKSCSCPTGCNSDDKCPCGNKSEETKKSCCSGK
13	Fungal 6	---	S.cerevisiae CRS5 TVKICDCEGECCKDSCHCGSTCLPSCSGGEKCKCDHSTGSPQCKSCGEKCKCETTCTCEKSKCNCEKC
14	Procaryota	K-C-A-C-x(2)-C-L-C	Synechococcus sp SmtA MTTVTQMKCACPHCLCIVSLNDAIMVDGKPYCSEVCANGTCKENSGCGHAGCGCGSA
15	Plant
15.1	Plant MTs Type 1	C-X-C-X(3)- C-X-C-X(3)- C-X-C-X(3)-spacer-C-X-C-X(3)- C-X-C-X(3)- C-X-C-X(3)	Pisum sativum MT MSGCGCGSSCNCGDSCKCNKRSSGLSYSEMETTETVILGVGPAKIQFEGAEMSAASEDGGCKCGDNCTCDPCNCK
15.2	Plant MTs Type 2	C-C-X(3)-C-X-C-X(3)- C-X-C-X(3)- C-X-C-X(3)-spacer- C-X-C-X(3)- C-X-C-X(3)- C-X-C-X(3)	L.esculetum MT MSCCGGNCGCGSSCKCGNGCGGCKMYPDMSYTESSTTTETLVLGVGPEKTSFGAMEMGESPVAENGCKCGSDCKCNPCTCSK
15.3	Plant MTs Type 3	---	A.thaliana MT3 MSSNCGSCDCADKTQCVKKGTSYTFDIVETQESYKEAMIMDVGAEENNANCKCKCGSSCSCVNCTCCPN
15.4	Plant MTs Type 4 or Ec	C-x(4)-C-X-C-X(3)-C-X(5)-C-X-C-X(9,11)-HTTCGCGEHC- X-C-X(20)-CSCGAXCNCASC-X(3,5)	T.aestium MT MGCNDKCGCAVPCPGGTGCRCTSARSDAAAGEHTTCGCGEHCGCNPCACGREGTPSGRANRRANCSCGAACNCASCGSTTA
99	Phytochelatins and other non-proteinaceous MT-like polypeptides	---	S.pombe γEC-γEC-γECG

More data on this classification are discoverable at the Expasy metallothionein page.^[5]

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(II) or Cd(II) (the protein is folded so as to bind metals in two functionally independent domains, with a metallic cluster each), yeast and procariotyc MTs show a monodominial structure (one domain with a single metallic cluster). Although no structural data is available for molluscan, nematoda and Drosophila MTs, it is commonly assumed that the former are bidominial and the latter monodominial. No conclusive data are available for Plant MTs, but two possible structures have been proposed: 1) 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 (Zn(II), Cd(II),...), while yeast CUP1 is selective for monovalent metal ions (Cu(I), Ag(I),...). Strictly metal-selective MTs with metal-specific physiological functions were discovered by Dallinger et al. (1997) in pulmonate snails (Gastropoda, Mollusca).^[6] The Roman snail (Helix pomatia), for example, possesses a Cd-selective (CdMT) and a Cu-selective isoform (CuMT) involved in Cd detoxification and Cu regulation, respectively.^[6] 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 (Palacios et al. 2011).^[6][7]

A novel functional classification of MTs as Zn- or Cu-thioneins is currently being developed based on these functional preferences.

Yeast

Metallothioneins are characterised by an abundance of cysteine residues and a lack of generic secondary structure motifs. Yeast Metallothionein (MT) are also alternatively named, Copper metallothionein (CUP).

Function

This protein functions in primary metal storage, transport and detoxification.^[8] More specifically, Yeast MT stores copper so therefore protects the cell against copper toxicity by tightly chelating copper ions.

Structure

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.^[8]

Examples

Yeast MT can be found in the following:^[9]

• Saccharomyces cerevisiae
• Neurospora crassa

Function

Metal binding

Metallothionein has been documented to bind a wide range of metals including cadmium,^[10] zinc, mercury, copper, arsenic, silver, etc. Metallation of MT was previously reported to occur cooperatively but recent reports have provided strong evidence that metal-binding occurs via a sequential, noncooperative mechanism.^[11] The observation of partially metallated MT (that is, having some free metal binding capacity) 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(II) 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 (MTs) 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.^[12][13] Metallothionein also carries zinc ions (signals) from one part of the cell to another. When zinc enters a cell, it can be picked up by thionein (which thus becomes "metallothionein") and carried to another part of the cell where it is released to another organelle or protein. In this way the thionein-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.^[14] 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 oxidative stress has been confirmed by MT Knockout mutants, but some experiments propose also a prooxidant role for MTs.

Expression and regulation

Metallothionein gene expression is induced by a high variety of stimuli, as metal exposure, oxidative stress, glucocorticoids, hydric stress, etc. The level of the response to these inducers depends on the MT gene. MT genes present in their promotors specific sequences for the regulation of the expression, elements as metal response elements (MRE), glucocorticoid response elements (GRE), GC-rich boxes, basal level elements (BLE), and thyroid response elements (TRE).

Metallothionein and disease

Cancer

Because MTs play an important role in transcription factor regulation, problems with MT function or expression may lead to malignant transformation of cells and ultimately cancer.^[15] Studies have found increased expression of MTs in some cancers of the breast, colon, kidney, liver, skin (melanoma), 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.^[16]

There is evidence to suggest that higher levels of MT expression may also lead to resistance to chemotherapeutic drugs. ^[17]

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.^[18] 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.^[19]

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.^[20]

And further there is indication that the mothers zinc levels may affect the developing babies immunological state that may lead to autism and could be again an indication that the Metallothionein system has been affected ^[21]

See also

• Phytochelatin

References

1. PDB: 2KAK​; Peroza EA, Schmucki R, Güntert P, Freisinger E, Zerbe O (March 2009). "The beta(E)-domain of wheat E(c)-1 metallothionein: a metal-binding domain with a distinctive structure". J. Mol. Biol. 387 (1): 207–18. doi:10.1016/j.jmb.2009.01.035. PMID 19361445.
2. Sigel H, Sigel A, eds. (2009). Metallothioneins and Related Chelators (Metal Ions in Life Sciences). Metal Ions in Life Sciences 5. Cambridge, England: Royal Society of Chemistry. ISBN 1-84755-899-2.
3. Margoshes M, Vallee BL (1957). "A cadmium protein from equine kidney cortex". Journal of the American Chemical Society 79 (17): 4813–4814. doi:10.1021/ja01574a064.
4. Felizola SJ, Nakamura Y, Arata Y, Ise K, Satoh F, Rainey WE, Midorikawa S, Suzuki S, Sasano H (2014). "Metallothionein-3 (MT-3) in the human adrenal cortex and its disorders". Endocrine Pathology 25 (3): 229–235. doi:10.1007/s12022-013-9280-9. PMID 24242700.
5. "Metallothioneins: classification and list of entries". www.uniprot.org.
6. Dallinger R, Berger B, Hunziker P, Kägi JHR (1997). "Metallothionein in snail Cd and Cu metabolism". Nature 388 (6639): 237–238.
7. Palacios Ò, Pagani A, Pérez-Rafael S, Egg M, Höckner M, Brandstätter A, Capdevila M, Atrian S, Dallinger R (2011). "Shaping mechanisms of metal specificity in a family of metazoan metallothioneins: evolutionary differentiation of mollusc metallothioneins". BMC Biology 9 (4): 1–20.
8. Peterson CW, Narula SS, Armitage IM (January 1996). "3D solution structure of copper and silver-substituted yeast metallothioneins". FEBS Lett. 379 (1): 85–93. doi:10.1016/0014-5793(95)01492-6. PMID 8566237.
9. Butt TR, Ecker DJ (September 1987). "Yeast metallothionein and applications in biotechnology". Microbiol. Rev. 51 (3): 351–64. PMC 373116. PMID 3312986.
10. Freisinger E, Vašák M (2013). "Cadmium in metallothioneins". Met Ions Life Sci 11: 339–71. doi:10.1007/978-94-007-5179-8_11. PMID 23430778.
11. Krezel A, Maret W (September 2007). "Dual nanomolar and picomolar Zn(II) binding properties of metallothionein". J. Am. Chem. Soc. 129 (35): 10911–21. doi:10.1021/ja071979s. PMID 17696343.
12. Huang M, Krepkiy D, Hu W, and Petering D (2004). "Zn-, Cd-, and Pb-transcription factor IIIA: properties, DNA binding, and comparison with TFIIIA-finger 3 metal complexes". Journal of Inorganic Biochemistry 98 (5): 775–785. doi:10.1016/j.jinorgbio.2004.01.014. PMID 15134923.
13. Huang M, Shaw CF, and Petering D (2004). "Interprotein metal exchange between transcription factor IIIa and apo-metallothionein". Journal of Inorganic Biochemistry 98 (4): 639–648. doi:10.1016/j.jinorgbio.2004.02.004. PMID 15041244.
14. Kumari MV, Hiramatsu M, Ebadi M (August 1998). "Free radical scavenging actions of metallothionein isoforms I and II". Free Radic. Res. 29 (2): 93–101. doi:10.1080/10715769800300111. PMID 9790511.
15. Krizkova S, Fabrik I, Adam V, Hrabeta J, Eckschlager T, Kizek R (2009). "Metallothionein--a promising tool for cancer diagnostics". Bratisl Lek Listy 110 (2): 93–7. PMID 19408840.
16. Cherian, M. (2003). "Metallothioneins in human tumors and potential roles in carcinogenesis". Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis 533: 201–209. doi:10.1016/j.mrfmmm.2003.07.013. PMID 14643421.
17. Basu, A. (2010). "Cellular Responses to Cisplatin-Induced DNA Damage". Journal of Nucleic Acids 2010. doi:10.4061/2010/201367.
18. Drum DA (October 2009). "Are toxic biometals destroying your children's future?". Biometals 22 (5): 697–700. doi:10.1007/s10534-009-9212-9. PMID 19205900.
19. Singh VK, Hanson J (June 2006). "Assessment of metallothionein and antibodies to metallothionein in normal and autistic children having exposure to vaccine-derived thimerosal". Pediatr Allergy Immunol 17 (4): 291–6. doi:10.1111/j.1399-3038.2005.00348.x. PMID 16771783.
20. Faber, S (2009). "The plasma zinc/serum copper ratio as a biomarker in children". Biomarkers 14: 171–80. doi:10.1080/13547500902783747. PMID 19280374.
21. Guillermo, Vela (2015). "Zinc in Gut-Brain Interaction in Autism". Neural Plast. 2015: 972791. doi:10.1155/2015/972791. PMC 4386645. PMID 25878905.

Further reading

• Cherian MG, Jayasurya A, Bay BH (December 2003). "Metallothioneins in human tumors and potential roles in carcinogenesis". Mutat. Res. 533 (1–2): 201–9. doi:10.1016/j.mrfmmm.2003.07.013. PMID 14643421.

External links

• Expasy metallothionein page