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1. Activity of Cytosolic Ascorbate Peroxidase (APX) from Panicum virgatum against Ascorbate and Phenylpropanoids

   https://doi.org/10.3390/ijms24021778  

   Zhang, Bixia; Lewis, Jacob A.; Kovacs, Frank; Sattler, Scott E.; Sarath, Gautam; Kang, ChulHee (January 2023, International Journal of Molecular Sciences)

   APX is a key antioxidant enzyme in higher plants, scavenging H2O2 with ascorbate in several cellular compartments. Here, we report the crystal structures of cytosolic ascorbate peroxidase from switchgrass (Panicum virgatum L., Pvi), a strategic feedstock plant with several end uses. The overall structure of PviAPX was similar to the structures of other APX family members, with a bound ascorbate molecule at the ɣ-heme edge pocket as in other APXs. Our results indicated that the H2O2-dependent oxidation of ascorbate displayed positive cooperativity. Significantly, our study suggested that PviAPX can oxidize a broad range of phenylpropanoids with δ-meso site in a rather similar efficiency, which reflects its role in the fortification of cell walls in response to insect feeding. Based on detailed structural and kinetic analyses and molecular docking, as well as that of closely related APX enzymes, the critical residues in each substrate-binding site of PviAPX are proposed. Taken together, these observations shed new light on the function and catalysis of PviAPX, and potentially benefit efforts improve plant health and biomass quality in bioenergy and forage crops. 

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2. A sorghum ascorbate peroxidase with four binding sites has activity against ascorbate and phenylpropanoids

   https://doi.org/10.1093/plphys/kiac604  

   Zhang, Bixia; Lewis, Jacob A; Vermerris, Wilfred; Sattler, Scott E; Kang, ChulHee (December 2022, Plant Physiology)

   Abstract In planta, H2O2 is produced as a by-product of enzymatic reactions and during defense responses. Ascorbate peroxidase (APX) is a key enzyme involved in scavenging cytotoxic H2O2. Here, we report the crystal structure of cytosolic APX from sorghum (Sorghum bicolor) (Sobic.001G410200). While the overall structure of SbAPX was similar to that of other APXs, SbAPX uniquely displayed four bound ascorbates rather than one. In addition to the ɣ-heme pocket identified in other APXs, ascorbates were bound at the δ-meso and two solvent-exposed pockets. Consistent with the presence of multiple binding sites, our results indicated that the H2O2-dependent oxidation of ascorbate displayed positive cooperativity. Bound ascorbate at two surface sites established an intricate proton network with ascorbate at the ɣ-heme edge and δ-meso sites. Based on crystal structures, steady-state kinetics, and site-directed mutagenesis results, both ascorbate molecules at the ɣ-heme edge and the one at the surface are expected to participate in the oxidation reaction. We provide evidence that the H2O2-dependent oxidation of ascorbate by APX produces a C2-hydrated bicyclic hemiketal form of dehydroascorbic acid at the ɣ-heme edge, indicating two successive electron transfers from a single-bound ascorbate. In addition, the δ-meso site was shared with several organic compounds, including p-coumaric acid and other phenylpropanoids, for the potential radicalization reaction. Site-directed mutagenesis of the critical residue at the ɣ-heme edge (R172A) only partially reduced polymerization activity. Thus, APX removes stress-generated H2O2 with ascorbates, and also uses this same H2O2 to potentially fortify cell walls via oxidative polymerization of phenylpropanoids in response to stress. 

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3. ASCORBATE PEROXIDASE6 delays the onset of age-dependent leaf senescence

   https://doi.org/10.1093/plphys/kiaa031  

   Chen, Changming; Galon, Yael; Rahmati Ishka, Maryam; Malihi, Shimrit; Shimanovsky, Vladislava; Twito, Shir; Rath, Abhishek; Vatamaniuk, Olena K; Miller, Gad (November 2020, Plant Physiology)

   null (Ed.)

   Abstract Age-dependent changes in reactive oxygen species (ROS) levels are critical in leaf senescence. While H2O2-reducing enzymes such as catalases and cytosolic ASCORBATE PEROXIDASE1 (APX1) tightly control the oxidative load during senescence, their regulation and function are not specific to senescence. Previously, we identified the role of ASCORBATE PEROXIDASE6 (APX6) during seed maturation in Arabidopsis (Arabidopsis thaliana). Here, we show that APX6 is a bona fide senescence-associated gene. APX6 expression is specifically induced in aging leaves and in response to senescence-promoting stimuli such as abscisic acid (ABA), extended darkness, and osmotic stress. apx6 mutants showed early developmental senescence and increased sensitivity to dark stress. Reduced APX activity, increased H2O2 level, and altered redox state of the ascorbate pool in mature pre-senescing green leaves of the apx6 mutants correlated with the early onset of senescence. Using transient expression assays in Nicotiana benthamiana leaves, we unraveled the age-dependent post-transcriptional regulation of APX6. We then identified the coding sequence of APX6 as a potential target of miR398, which is a key regulator of copper redistribution. Furthermore, we showed that mutants of SQUAMOSA PROMOTER BINDING PROTEIN-LIKE7 (SPL7), the master regulator of copper homeostasis and miR398 expression, have a higher APX6 level compared with the wild type, which further increased under copper deficiency. Our study suggests that APX6 is a modulator of ROS/redox homeostasis and signaling in aging leaves that plays an important role in developmental- and stress-induced senescence programs. 

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4. Cyclic electron flow (CEF) and ascorbate pathway activity provide constitutive photoprotection for the photopsychrophile, Chlamydomonas sp. UWO 241 (renamed Chlamydomonas priscuii)

   https://doi.org/10.1007/s11120-021-00877-5  

   Stahl-Rommel, Sarah; Kalra, Isha; D’Silva, Susanna; Hahn, Mark M.; Popson, Devon; Cvetkovska, Marina; Morgan-Kiss, Rachael M. (March 2022, Photosynthesis Research)
5. Completely Multipolar Model for Many-Body Water–Ion and Ion–Ion Interactions

   https://doi.org/10.1021/acs.jpclett.4c02940  

   Heindel, Joseph P; Kim, Lukas; Head-Gordon, Martin; Head-Gordon, Teresa (January 2025, The Journal of Physical Chemistry Letters)