Independent metal-binding features of recombinant metallothioneins convergently draw a step gradation between Zn- and Cu-thioneins

Bofill, Roger; Capdevila, Mercè; Atrian, Sı́lvia

doi:10.1039/b904953c

Cited by 67 publications

(97 citation statements)

References 24 publications

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“…4b). The identification of this sequence as a CaCuMT isoform was corroborated by the fact that, upon recombinant synthesis in E. coli of the corresponding cDNA , the resulting protein was a unique, homometallic Cu-loaded complex, a behaviour exhibited only by highly specific Cu-thioneins (Bofill et al 2009). This isoform was equally detected in Cd-fed snails-without major changes in its abundance, elution time, Cu load, and Cu:Cd:Zn molar ratio (Fig.…”

Section: Transcription Of the Cacd/cumt Genementioning

confidence: 89%

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Physiological relevance and contribution to metal balance of specific and non-specific Metallothionein isoforms in the garden snail, Cantareus aspersus

et al. 2011

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Variable environmental availability of metal ions represents a constant challenge for most organisms, so that during evolution, they have optimised physiological and molecular mechanisms to cope with this particular requirement. Metallothioneins (MTs) are proteins that play a major role in metal homeostasis and as a reservoir. The MT gene/protein systems of terrestrial helicid snails are an invaluable model for the study of metal-binding features and MT isoform-specific functionality of these proteins. In the present study, we characterised three paralogous MT isogenes and their expressed products in the escargot (Cantareus aspersus). The metal-dependent transcriptional activation of the three isogenes was assessed using quantitative Real Time PCR. The metal-binding capacities of the three isoforms were studied by characterising the purified native complexes. All the data were analysed in relation to the trace element status of the animals after metal feeding. Two of the three C. aspersus MT (CaMT) isoforms appeared to be metal-specific, (CaCdMT and CaCuMT, for cadmium and copper respectively). A third isoform (CaCd/CuMT) was non-specific, since it was natively recovered as a mixed Cd/Cu complex. A specific role in Cd detoxification for CaCdMT was revealed, with a 80-90% contribution to the Cd balance in snails exposed to this metal. Conclusive data were also obtained for the CaCuMT isoform, which is involved in Cu homeostasis, sharing about 30-50% of the Cu balance of C. aspersus. No apparent metal-related physiological function was found for the third isoform (CaCd/CuMT), so its contribution to the metal balance of the escargot may be, if at all, of only marginal significance, but may enclose a major interest in evolutionary studies.

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Section: Transcription Of the Cacd/cumt Genementioning

confidence: 89%

“…Furthermore, some of the Cd-species include S 2-ligands (Fig. 5c), which is a trait indicative of Cu-thionein character (Capdevila et al 2005;Bofill et al 2009;Orihuela et al 2010). Concordantly, this isoform exhibits intermediate features regarding its induction pattern and synthesis.…”

Section: Specificity Of the Metal-binding Behaviour Of The Camt Isoformsmentioning

confidence: 92%

Physiological relevance and contribution to metal balance of specific and non-specific Metallothionein isoforms in the garden snail, Cantareus aspersus

et al. 2011

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“…It is now well established that, in general, the ESI-MS process is brief enough to indeed detect proteins with essentially natively folded structures [89]. With regard to metalloproteins (a dedicated review is available elsewhere [90]), many studies concern Ca-binding proteins, for example calmodulin [91], heme-binding proteins [92][93][94], zinc fingers [95][96][97], and metallothioneins [97][98][99][100][101]. The vast majority of these studies used ESI-MS rather than MALDI-MS, although early studies on copper binding to small peptides used both ionisation methods [102].…”

Section: Immobilised Metal-affinity Chromatographymentioning

confidence: 99%

Protein fractionation and detection for metalloproteomics: challenges and approaches

2012

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At least one third of all proteins are thought to require a metal ion co-factor for their function. Recognition of the importance of metals in biological systems and major advances in analytical instrumentation and technology have led to the emergence of the new research area of metalloproteomics in recent years. Despite this progress, the experimental determination of in-vivo metal cofactors has remained challenging, because this requires elucidation of protein interactions with non-covalently bound metal ions. This critical review highlights current methodological approaches, focusing, in particular, on issues relating to the fractionation and separation of the metalloproteome, including recent experience with metalloproteomics for marine cyanobacteria in our laboratory. Metalloproteomics promises to deliver novel insights into fundamental biological processes in the future, but it is clear that further methodological advances are necessary to exploit the full potential of this emerging research area.

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“…It is well documented that different MTs exhibit distinct preferences for binding either divalent or monovalent metal ions, which allowed their classification as extreme Zn-or Cu-thioneins, or intermediate MTs in a gradation of continuum metal-binding behaviours [2,41]. Genuine Zn-thioneins are MTs optimized for divalent metal binding, which render unique species when synthesized as Zn-MT or Cd-complexes and several mixed Zn, Cu-MT complexes of different stoichiometries when biosynthesized in a Cu-rich environment.…”

Section: Mass Spectrometry Circular Dichroism and Uv-vis Analysismentioning

confidence: 99%

Rhenium and technetium tricarbonyl, {M(CO)3}+ (M = Tc, Re), binding to mammalian metallothioneins: new insights into chemical and radiopharmaceutical implications

et al. 2014

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This paper deals with the binding of the four mammalian metallothioneins (MTs) to the organometallic metal fragment {fac-M(CO)3}(+) (M = (99)Tc, Re), which is highly promising for the preparation of second-generation radiopharmaceuticals. The study of the transmetallation reaction between zinc and rhenium in Zn7-MT1 by means of UV-vis and CD spectroscopy demonstrated the incorporation of the {fac-Re(CO)3}(+) fragment to the MTs. This reaction should be performed at 70 °C to accelerate the reaction rate, a result that is consistent with the reported reactivity of the rhenium fragment. ESI-TOF MS demonstrated the formation of mixed-metal species as Zn6,{Re(CO)3}-MT, Zn6,{Re(CO)3}2-MT, and Zn5,{Re(CO)3}3-MT, as well as the different reactivity of the four MT isoforms. Hence, Zn-MT3 showed the highest reactivity, in agreement with its high Cu-thionein character, whereas Zn-MT2 exhibited the lowest reactivity, in line with its high Zn-thionein character. The reactivity of the Zn-loaded forms of MT1 and MT4 is intermediate between those of MT3 and MT2. The study of the binding of the {fac-(99)Tc(CO)3}(+) fragment to MTs showed a significant and very interesting different reactivity in relation to rhenium. The transmetallation reaction is much more effective with technetium than with rhenium and significant amounts of mixed Zn x ,{(99)Tc(CO)3} y -MT species were formed with the four MT isoforms whereas only MT3 rendered similar amounts of rhenium derivatives. The results obtained in this study support the possible use of technetium for labelling mammalian metallothioneins and also for possible radiopharmaceutical applications.

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Independent metal-binding features of recombinant metallothioneins convergently draw a step gradation between Zn- and Cu-thioneins

Cited by 67 publications

References 24 publications

Physiological relevance and contribution to metal balance of specific and non-specific Metallothionein isoforms in the garden snail, Cantareus aspersus

Physiological relevance and contribution to metal balance of specific and non-specific Metallothionein isoforms in the garden snail, Cantareus aspersus

Protein fractionation and detection for metalloproteomics: challenges and approaches

Rhenium and technetium tricarbonyl, {M(CO)3}+ (M = Tc, Re), binding to mammalian metallothioneins: new insights into chemical and radiopharmaceutical implications

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