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Characterization analysis and heavy metal-binding properties of CsMTL3 in Escherichia coli.

ABSTRACT: Members of the metallothionein (MT) superfamily are involved in coordinating transition metal ions. In plants, MT family members are characterized by their arrangement of Cys residues. In this study, one member of the MT superfamily, CsMTL3, was characterized from a complementary DNA (cDNA) library from young cucumber fruit; CsMTL3 is predicted to encode a 64 amino acid protein with a predicted molecular mass of 6.751 kDa. Phylogenetic analysis identified it as a type 3 family member as the arrangement of N-terminal Cys residues was different from that of MT-like 2. Heterologous expression of CsMTL3 in Escherichia coli improved their heavy metal tolerance, particularly to Cd2+ and Cu2+, and led to increased uptake of Cd2+ and Cu2+; increased uptake was also observed for cells expressing Arabidopsis thaliana metallothionein 3 (AtMT3) and phytochelatin-like (PCL), with greatest uptake in PCL-expressing cells. These findings demonstrate that CsMTL3 can improve metal tolerance, especially for Cd2+ ions, when heterologously expressed in E. coli, and suggest that the composition and arrangement of N-terminal Cys residues are associated with binding capacity and preference for different metal ions.

SUBMITTER: Xu X

PROVIDER: S-EPMC6212650 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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Characterization analysis and heavy metal-binding properties of CsMTL3 in Escherichia coli.

Xu Xing X Duan Ling L Yu Jingwen J Su Chenggang C Li Jinhua J Chen Dan D Zhang Xingguo X Song Hongyuan H Pan Yu Y

FEBS open bio 20180919 11

Members of the metallothionein (MT) superfamily are involved in coordinating transition metal ions. In plants, MT family members are characterized by their arrangement of Cys residues. In this study, one member of the MT superfamily, CsMTL3, was characterized from a complementary DNA (cDNA) library from young cucumber fruit; CsMTL3 is predicted to encode a 64 amino acid protein with a predicted molecular mass of 6.751 kDa. Phylogenetic analysis identified it as a type 3 family member as t ...[more]

PMID: 30410861

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Project description:In Escherichia coli and other bacteria, nickel uptake is regulated by the transcription factor NikR. Nickel binding at high-affinity sites in E. coli NikR (EcNikR) facilitates EcNikR binding to the nik operon, where it then suppresses transcription of genes encoding the nickel uptake transporter, NikABCDE. A structure of the EcNikR-DNA complex suggests that a second metal-binding site is also present when NikR binds to the nik operon. Moreover, this co-crystal structure raises the question of what metal occupies the second site under physiological conditions: K(+), which is present in the crystal structure, or Ni(2+), which has been proposed to bind to low- as well as high-affinity sites on EcNikR. To determine which ion is preferred at the second metal-binding site and the physical basis for any preference of one ion over another in both the second metal-binding site and the high-affinity sites, we conducted a series of detailed molecular simulations on the EcNikR structure. Simulations that place Ni(2+) at high-affinity sites lead to stable trajectories with realistic ion-ligand distances and geometries, while simulations that place K(+) at these sites lead to conformational changes in the protein that are likely unfavorable for ion binding. By contrast, simulations on the second metal site in the EcNikR-DNA complex lead to stable trajectories with realistic geometries regardless of whether K(+) or Ni(2+) occupies this site. Electrostatic binding free energy calculations, however, suggest that EcNikR binding to DNA is more favorable when the second metal-binding site contains K(+). An analysis of the energetic contributions to the electrostatic binding free energy suggests that, while the interaction between EcNikR and DNA is more favorable when the second site contains Ni(2+), the large desolvation penalty associated with moving Ni(2+) from solution to the relatively buried second site offsets this favorable interaction term. Additional free energy simulations that account for both electrostatic and non-electrostatic effects argue that EcNikR binding to DNA is most favorable when the second site contains a monovalent ion the size of K(+). Taken together, these data suggest that the EcNikR structure is most stable when Ni(2+) occupies high-affinity sites and that EcNikR binding to DNA is more favorable when the second site contains K(+).

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Characterization analysis and heavy metal-binding properties of CsMTL3 in Escherichia coli.

Publications

Characterization analysis and heavy metal-binding properties of <i>CsMTL3</i> in <i>Escherichia coli</i>.

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Dataset Information

Characterization analysis and heavy metal-binding properties of CsMTL3 in Escherichia coli.

Publications

Characterization analysis and heavy metal-binding properties of &lt;i&gt;CsMTL3&lt;/i&gt; in &lt;i&gt;Escherichia coli&lt;/i&gt;.

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Characterization analysis and heavy metal-binding properties of <i>CsMTL3</i> in <i>Escherichia coli</i>.