Prashanth Kumar Koochana1, Abhinav Mohanty1, Suman Das1, Biswamaitree Subhadarshanee2, Suresh Satpati3, Anshuman Dixit3, Surendra Chandra Sabat3, Rabindra K Behera4. 1. Department of Chemistry, National Institute of Technology, Rourkela 769008, Odisha, India. 2. Department of Chemistry, National Institute of Technology, Rourkela 769008, Odisha, India; KIIT School of Biotechnology, KIIT University, Bhubaneswar 751024, Odisha, India. 3. Institute of Life Sciences, Bhubaneswar 751023, Odisha, India. 4. Department of Chemistry, National Institute of Technology, Rourkela 769008, Odisha, India. Electronic address: beherarabi@nitrkl.ac.in.
Abstract
BACKGROUND: Ferritin detoxifies excess of free Fe(II) and concentrates it in the form of ferrihydrite (Fe2O3·xH2O) mineral. When in need, ferritin iron is released for cellular metabolic activities. However, the low solubility of Fe(III) at neutral pH, its encapsulation by stable protein nanocage and presence of dissolved O2 limits in vitro ferritin iron release. METHODS: Physiological reducing agent, NADH (E1/2 = -330 mV) was inefficient in releasing the ferritin iron (E1/2 = +183 mV), when used alone. Thus, current work investigates the role of low concentration (5-50 μM) of phenazine based electron transfer (ET) mediators such as FMN, PYO - a redox active virulence factor secreted by Pseudomonas aeruginosa and PMS towards iron mobilization from recombinant frog M ferritin. RESULTS: The presence of dissolved O2, resulting in initial lag phase and low iron release in FMN, had little impact in case of PMS and PYO, reflecting their better ET relay ability that facilitates iron mobilization. The molecular modeling as well as fluorescence studies provided further structural insight towards interaction of redox mediators on ferritin surface for electron relay. CONCLUSIONS: Reductive mobilization of iron from ferritin is dependent on the relative rate of NADH oxidation, dissolved O2 consumption and mineral core reduction, which in turn depends on E1/2 of these mediators and their interaction with ferritin. GENERAL SIGNIFICANCE: The current mechanism of in vitro iron mobilization from ferritin by using redox mediators involves different ET steps, which may help to understand the iron release pathway in vivo and to check microbial growth.
BACKGROUND: Ferritin detoxifies excess of free Fe(II) and concentrates it in the form of ferrihydrite (Fe2O3·xH2O) mineral. When in need, ferritin iron is released for cellular metabolic activities. However, the low solubility of Fe(III) at neutral pH, its encapsulation by stable protein nanocage and presence of dissolved O2 limits in vitro ferritin iron release. METHODS: Physiological reducing agent, NADH (E1/2 = -330 mV) was inefficient in releasing the ferritin iron (E1/2 = +183 mV), when used alone. Thus, current work investigates the role of low concentration (5-50 μM) of phenazine based electron transfer (ET) mediators such as FMN, PYO - a redox active virulence factor secreted by Pseudomonas aeruginosa and PMS towards iron mobilization from recombinant frog M ferritin. RESULTS: The presence of dissolved O2, resulting in initial lag phase and low iron release in FMN, had little impact in case of PMS and PYO, reflecting their better ET relay ability that facilitates iron mobilization. The molecular modeling as well as fluorescence studies provided further structural insight towards interaction of redox mediators on ferritin surface for electron relay. CONCLUSIONS: Reductive mobilization of iron from ferritin is dependent on the relative rate of NADH oxidation, dissolved O2 consumption and mineral core reduction, which in turn depends on E1/2 of these mediators and their interaction with ferritin. GENERAL SIGNIFICANCE: The current mechanism of in vitro iron mobilization from ferritin by using redox mediators involves different ET steps, which may help to understand the iron release pathway in vivo and to check microbial growth.