A metallothionein type 2 from Avicennia marina binds to iron and mediates hydrogen peroxide balance by activation of enzyme catalase. (August 2020)

• Record Type: Journal Article
• Authors: Babaei-Bondarti, Zahra; Shahpiri, Azar
• Is Part Of: Phytochemistry. Volume 176(2020)
• Publication Date: 2020-08
• Subjects: Avicennia marina -- Acanthaceae -- Metallothionein -- Metal accumulation -- Recombinant protein -- Iron -- Hydrogen peroxide tolerance; AmMT Avicennia marina MT -- CAT Catalase -- GST, DTNB 5 5′-dithio-bis-[2-nitrobenzoic acid] -- GST Glutathion-S-transferase -- IPTG Isopropyl β-D-1-thiogalactopyranoside -- LB Luria-bertani -- MTs Metallothioneins -- PCs Phytochelatins -- ROS Reactive Oxygen species; Botanical chemistry -- Periodicals; Biochemistry -- Periodicals; Botany -- Periodicals; Chimie végétale -- Périodiques; 572.2
• DOI: 10.1016/j.phytochem.2020.112396
• Languages: English
• ISSNs: 0031-9422

Abstract

Abstract: Metallothioneins (MTs) are low molecular weight, cysteine-rich, metal-binding proteins that are important for essential metal homeostasis, protection against oxidative stress, and buffering against toxic heavy metals. In this work the gene encoding an MT type 2 from Avicennia marina (Forssk.) Vierh. (AmMT2) was cloned into pET41a and transformed into the Escherichia coli strain Rosetta (DE3). Following the induction with isopropyl β-D -1-thiogalactopyranoside, AmMT2 was expressed as glutathione-S-transferase (GST)-tagged fusion protein. The accumulation of Zn 2+, Cu 2+, Fe 2+, Ni 2+ and Cd 2+ for strain R-AmMT2 was 4, 8, 5.4, 2 and 1.6 fold of control strain suggesting the role of AmMT2 in accumulation of metals. Particularly the strain R-AmMT2 was able to accumulate 30.7 mg per g dry weight. The cells expressing AmMT2 was more tolerant to hydrogen peroxide and had higher catalase (CAT) activity. To understand the mechanistic action of AmMT2 hydrogen peroxide tolerance, the activity of CAT in the E. coli protein extract was assayed after addition of pure Fe 2+ /GST-AmMT complex and Apo/GST-AmMT2 in vitro . Whereas, the activity of CAT did not change by the addition of Apo/GST-AmMT2, the activity of CAT significantly increased after addition of Fe 2+ /GST-AmMT2. These results show that AmMT2 activates CAT through Fe 2+ transfer which subsequently causes the oxidative stress tolerance. Graphical abstract: Image 1 Highlights: The transgenic E. coli expressing AmMT2 removed the significant amounts of Zn 2+, Cu 2+, Fe 2+ and Ni 2 . Transgenic strain expressing GST-AmMT2 was able to accumulate 30.7 mg Fe 2+ per gr (dry weight cells). The pure and apo-form of GST-AmMT2 was able to bind to Fe 2+ in vitro. The activity of catalase in R-AmMT2 was significantly higher than control strain. The protein Fe 2+ /AmMT2 seems to enhance the activity of catalase by providing Fe 2+ ion for this enzyme.