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Heterologous Expression and Metal-Binding Characterization of a Type 1 Metallothionein Isoform (OsMTI-1b) from Rice (Oryza sativa)

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Abstract

Metallothioneins (MTs) are ubiquitous, low molecular mass and cysteine-rich proteins that play important roles in maintaining intracellular metal homeostasis, eliminating metal toxification and protecting the cells against oxidative damages. MTs are able to bind metal ions through the thiol groups of their cysteine residues. Plants have several MT isoforms which are classified into four types based on the arrangement of cysteine residues. In the present study, a rice (Oryza sativa) gene encoding type 1 MT isoform, OsMTI-1b, was inserted in vector pET41a and overexpressed in Escherichia coli as carboxy-terminal extensions of glutathione-S-transferase (GST). The recombinant protein GST-OsMTI-1b was purified using affinity chromatography and its ability to bind with Ni2+, Cd2+, Zn2+ and Cu2+ ions was analyzed. The results demonstrated that this isoform has ability to bind Ni2+, Cd2+ and Zn2+ ions in vitro, whereas it has no substantial ability to bind Cu2+ ions. From competitive reaction with 5,5′-dithiobis(2-nitrobenzoic acid), DTNB, the affinity of metal ions for recombinant form of GST-OsMTI-1b was as follows: Ni2+/Cd2+ > Zn2+ > Cu2+

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Abbreviations

DTNB:

5,5′-Dithiobis(2-nitrobenzoic) acid

GST:

Glutathione-S-transferase

IPTG:

Isopropyl β-d-thiogalactoside

MT:

Metallothionein

Os:

Oryza sativa

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Acknowledgments

Iran National Science Foundation is thanked for financial support.

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Authors and Affiliations

  1. Department of Agriculture Biotechnology, College of Agriculture, Isfahan University of Technology, 84156-83111, Isfahan, Iran

    Rezvan Mohammadi Nezhad, Azar Shahpiri & Aghafakhr Mirlohi

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  1. Rezvan Mohammadi Nezhad
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  2. Azar Shahpiri
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  3. Aghafakhr Mirlohi
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Correspondence to Azar Shahpiri.

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Nezhad, R.M., Shahpiri, A. & Mirlohi, A. Heterologous Expression and Metal-Binding Characterization of a Type 1 Metallothionein Isoform (OsMTI-1b) from Rice (Oryza sativa). Protein J 32, 131–137 (2013). https://doi.org/10.1007/s10930-013-9469-2

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