title: Functional characterization of a type 3 metallolthionein isoform (OsMTI-3a) from rice.
abstract: Metallothioneins (MTs) are low-molecular weight proteins with high Cys content and a high affinity for metals. Plant MTs are classified into four types based on the arrangement of Cys in their amino acid sequences. In the present study, the gene encoding OsMTI-3a, a type 3 MT found in rice, was cloned into pET41a vector. The resulting construct was transformed into the Escherichia coli strain Rosetta (DE3). Following the induction with isopropyl β-D-1-thiogalactopyranoside, the OsMTI-3a was expressed as glutathione-S-transferase (GST)-tagged fusion protein. In comparison to control strain, the cells expressing GST-OsMTI-3a accumulated more Cd(2+), Ni(2+) and Zn(2+) when they were grown in the medium containing CdCl2, NiCl2 or ZnSO4. The recombinant GST-OsMTI-3a was purified using affinity chromatography. The UV absorption spectra recorded after the reconstitution of the apo-protein with different metals confirmed that GST-OsMTI-3a was able to form complexes with Cd(2+), Ni(2+), and Zn(2+). The reaction of the protein-metal complexes with 5-5-dithiobis (2-nitrobenzoic) revealed that the order of affinity of GST-OsMTI-3a toward different metals was Ni(2+)≥Cd(2+)>Zn(2+)>Cu(2+).
authors.affiliation: Department of Agricultural Biotechnology, College of Agriculture, Isfahan University of Technology, Isfahan 84156-83111, Iran. Electronic address: a.shahpiri@cc.iut.ac.ir.
authors.forename: Azar
authors.initials: A
authors.lastname: Shahpiri
authors.affiliation: Department of Agricultural Biotechnology, College of Agriculture, Isfahan University of Technology, Isfahan 84156-83111, Iran.
authors.forename: Iman
authors.initials: I
authors.lastname: Soleimanifard
authors.affiliation: Department of Biotechnology, Faculty of Advanced Sciences and Technologies, University of Isfahan, Isfahan 81746-73441, Iran.
authors.forename: Mohammad Ali
authors.initials: MA
authors.lastname: Asadollahi
mesh_terms: D015870:Gene Expression; D023061:Gene Order; D005822:Genetic Vectors / Q000235:genetics; D007477:Ions / Q000737:chemistry / Q000378:metabolism; D008668:Metallothionein / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008670:Metals / Q000737:chemistry / Q000378:metabolism; D012275:Oryza / Q000737:chemistry* / Q000235:genetics; D011485:Protein Binding; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D007477:Ions; D008670:Metals; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; D008668:Metallothionein
keywords: Functional characterization; Heavy metals; Heterologous expression; Metallothionein; Rice