TY - JOUR
T1 - Ascorbate Peroxidase 2 (APX2) of Chlamydomonas Binds Copper and Modulates the Copper Insertion into Plastocyanin
AU - Caccamo, Anna
AU - Vega de Luna, Félix
AU - Wahni, Khadija
AU - Volkov, A.N.
AU - Przybyla-Toscano, Jonathan
AU - Amelii, Antonello
AU - Kriznik, Alexandre
AU - Rouhier, Nicolas
AU - Messens, Joris
AU - Remacle, Claire
N1 - Funding Information:
This work was funded by Fonds de la Recherche Scientifique-the Research Foundation Flanders—Excellence of Science project No. 30829584 (to C.R. and J.M.) and a VIB-grant (to J.M.); A.C. acknowledges a PhD fellowship of ULiège supported by Fonds de la Recherche Scientifique Excellence of Science project No. 30829584; J.P.-T. is postdoc from Belgian Fonds de la Recherche Scientifique-FNRS, respectively. The Hubert Curien Tournesol partnership is acknowledged by N.R. and C.R.
Publisher Copyright:
© 2023 by the authors.
PY - 2023/11
Y1 - 2023/11
N2 - Recent phylogenetic studies have unveiled a novel class of ascorbate peroxidases called "ascorbate peroxidase-related" (APX-R). These enzymes, found in green photosynthetic eukaryotes, lack the amino acids necessary for ascorbate binding. This study focuses on the sole APX-R from Chlamydomonas reinhardtii referred to as ascorbate peroxidase 2 (APX2). We used immunoblotting to locate APX2 within the chloroplasts and in silico analysis to identify key structural motifs, such as the twin-arginine transport (TAT) motif for lumen translocation and the metal-binding MxxM motif. We also successfully expressed recombinant APX2 in Escherichia coli. Our in vitro results showed that the peroxidase activity of APX2 was detected with guaiacol but not with ascorbate as an electron donor. Furthermore, APX2 can bind both copper and heme, as evidenced by spectroscopic, and fluorescence experiments. These findings suggest a potential interaction between APX2 and plastocyanin, the primary copper-containing enzyme within the thylakoid lumen of the chloroplasts. Predictions from structural models and evidence from 1H-NMR experiments suggest a potential interaction between APX2 and plastocyanin, emphasizing the influence of APX2 on the copper-binding abilities of plastocyanin. In summary, our results propose a significant role for APX2 as a regulator in copper transfer to plastocyanin. This study sheds light on the unique properties of APX-R enzymes and their potential contributions to the complex processes of photosynthesis in green algae.
AB - Recent phylogenetic studies have unveiled a novel class of ascorbate peroxidases called "ascorbate peroxidase-related" (APX-R). These enzymes, found in green photosynthetic eukaryotes, lack the amino acids necessary for ascorbate binding. This study focuses on the sole APX-R from Chlamydomonas reinhardtii referred to as ascorbate peroxidase 2 (APX2). We used immunoblotting to locate APX2 within the chloroplasts and in silico analysis to identify key structural motifs, such as the twin-arginine transport (TAT) motif for lumen translocation and the metal-binding MxxM motif. We also successfully expressed recombinant APX2 in Escherichia coli. Our in vitro results showed that the peroxidase activity of APX2 was detected with guaiacol but not with ascorbate as an electron donor. Furthermore, APX2 can bind both copper and heme, as evidenced by spectroscopic, and fluorescence experiments. These findings suggest a potential interaction between APX2 and plastocyanin, the primary copper-containing enzyme within the thylakoid lumen of the chloroplasts. Predictions from structural models and evidence from 1H-NMR experiments suggest a potential interaction between APX2 and plastocyanin, emphasizing the influence of APX2 on the copper-binding abilities of plastocyanin. In summary, our results propose a significant role for APX2 as a regulator in copper transfer to plastocyanin. This study sheds light on the unique properties of APX-R enzymes and their potential contributions to the complex processes of photosynthesis in green algae.
KW - Chlamydomonas
KW - ascorbate peroxidase-related
KW - copper-binding motif
KW - green microalga
KW - peroxidase activity
KW - plastocyanin
KW - structural prediction
KW - 1H-NMR
UR - http://www.scopus.com/inward/record.url?scp=85178374544&partnerID=8YFLogxK
U2 - 10.3390/antiox12111946
DO - 10.3390/antiox12111946
M3 - Article
VL - 12
JO - Antioxidants special issue on ‘Redox language of the cell'
JF - Antioxidants special issue on ‘Redox language of the cell'
SN - 1523-0864
IS - 11
M1 - 1946
ER -