Abstract
Leghemoglobin (Lb) is an oxygen-binding plant hemoglobin of legume nodules, which participates in the symbiotic nitrogen fixation process. Another way to obtain Lb is its expression in bacteria, yeasts, or other organisms. This is promising for both obtaining Lb in the necessary quantity and scrutinizing it in model systems, e.g., its interaction with reactive oxygen (ROS) and nitrogen (RNS) species. The main goal of the work was to study how Lb expression affected the ability of Escherichia coli cells to tolerate oxidative and nitrosative stress. The bacterium E. coli with the embedded gene of soybean leghemoglobin a contains this protein in an active oxygenated state. The interaction of the expressed Lb with oxidative and nitrosative stress inducers (nitrosoglutathione, tert-butyl hydroperoxide, and benzylviologen) was studied by enzymatic methods and spectrophotometry. Lb formed NO complexes with heme-nitrosylLb or nonheme iron-dinitrosyl iron complexes (DNICs). The formation of Lb-bound DNICs was also detected by low-temperature electron paramagnetic resonance spectroscopy. Lb displayed peroxidase activity and catalyzed the reduction of organic peroxides. Despite this, E. coli-synthesized Lb were more sensitive to stress inducers. This might be due to the energy demand required by the Lb synthesis, as an alien protein consumes bacterial resources and thereby decreases adaptive potential of E. coli.
Highlights
Leghemoglobin (Lb) is a symbiotic plant hemoglobin (Hb) synthesized in legume nodules that are formed on roots following their infection by nitrogen-fixing nodulating bacteria of several genera (Rhizobium, Bradyrhizobium, Azorhisobium, etc.)
The E. coli strain TB-1 used in our study had a high level of Lb expression, which was evidenced by the pronounced pink color of the cells grown at the liquid nutrient medium LB
Since Bv has the ability of single-electron oxidation and reduction, its biological activity is associated with electron transfer from the components of respiratory chain to oxygen with the formation of superoxide (O2−), which is a precursor of other reactive oxygen (ROS) and reactive nitrogen species (RNS), including hydrogen peroxide and peroxynitrite [41]
Summary
Leghemoglobin (Lb) is a symbiotic plant hemoglobin (Hb) synthesized in legume nodules that are formed on roots following their infection by nitrogen-fixing nodulating bacteria of several genera (Rhizobium, Bradyrhizobium, Azorhisobium, etc.). In nodules, these bacteria change their form and properties, turning into a symbiotic state bacteroids. The RNA sequencing analysis shows that Lb is expressed within a few hours after the inoculation and that its mRNA is found in roots and pods [10] This indicates that Lb functions are not confined to nitrogen fixation only
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