Redox control and autoxidation of class 1, 2 and 3 phytoglobins from Arabidopsis thaliana

Abstract

Despite a recent increase in interest towards phytoglobins and their importance in plants, much is still unknown regarding their biochemical/biophysical properties and physiological roles. The present study presents data on three recombinant Arabidopsis phytoglobins in terms of their UV-vis and Raman spectroscopic characteristics, redox state control, redox potentials and autoxidation rates. The latter are strongly influenced by pH for all three hemoglobins – (with a fundamental involvement of the distal histidine), as well as by added anion concentrations – suggesting either a process dominated by nucleophilic displacement of superoxide for AtHb2 or an inhibitory effect for AtHb1 and AtHb3. Reducing agents, such as ascorbate and glutathione, are found to either enhance– (presumably via direct electron transfer or via allosteric regulation) or prevent autoxidation. HbFe3+ reduction was possible in the presence of high (presumably not physiologically relevant) concentrations of NADH, glutathione and ascorbate, with differing behaviors for the three globins. The iron coordination sphere is found to affect the autoxidation, redox state interconversion and redox potentials in these three phytoglobins.

Plant hemoglobins: a journey from unicellular green algae to vascular plants

“…First, it lowers the midpoint redox potential for the protein compared to pentacoordinate Glbs because the His binds more tightly to Glb 3+ than to Glb 2+ , thus favoring the oxidized form of the protein. This has been observed in generally lower midpoint redox potentials for many of the plant Glbs (Halder et al , ; Mot et al , ). Second, it increases rate constants for electron transfer to and from the heme (Weiland et al , ) because there is minimal structural change in switching between oxidation states.…”

“…Regulation of ligand binding in support of function has been proposed by the detailed study of bimolecular and geminate ligand rebinding to Atha1 and Atha2 (Bruno et al , ; Spyrakis et al , ). Recently, extensive kinetic studies on the various Glbs of the model legume Lotus japonicus (Calvo‐Begueria et al , ) and Arabidopsis (Mot et al , ) strongly suggest non‐redundant functions.…”

Oxidative Implications of Substituting a Conserved Cysteine Residue in Sugar Beet Phytoglobin BvPgb 1.2

“…The dissociation of O 2 from the heme may ultimately lead to autoxidation, resulting in the formation of the ferric state (Fe III ) and superoxide (O 2 •− ) [ 16 ]. Thus, important insights regarding redox stability can be obtained by examining the autoxidation rates for the WT and the mutant.…”

“…In general, for Pgbs to act as a NO scavenger, the protein must bind O 2 first, followed by the incorporation of NO. However, the dissociation of O 2 from the heme may ultimately lead to autoxidation, resulting in forming the ferric state (Fe III ) and a superoxide radical (O 2 •− ) [ 16 ]. Thus, analysis of autoxidation rates are particularly relevant for globins because they are much less reactive in the ferric state and unable to bind O 2 , than in the ferrous (Fe II ) state [ 7 ].…”

Conditional Protein Function via N-Degron Pathway–Mediated Proteostasis in Stress Physiology

“…The effect was not as prominent for the other well-known hypoxia-responsive genes, ADH, PDC1, and PCOs (19). The three phytoglobins (plant hemoglobins) Hb1, Hb2, and Hb3 were shown only recently to have a strong tendency for autooxidation (91). Moreover, they act as NO scavengers via a NO-dioxygenase reaction and can bind O 2 (2,98).…”