Details
| Original language | English |
|---|---|
| Pages (from-to) | 917-934 |
| Number of pages | 18 |
| Journal | Journal of experimental botany |
| Volume | 75 |
| Issue number | 3 |
| Early online date | 16 Oct 2023 |
| Publication status | Published - 2 Feb 2024 |
Abstract
Proline dehydrogenase (ProDH) and pyrroline-5-carboxylate (P5C) dehydrogenase (P5CDH) catalyze the oxidation of proline into glutamate via the intermediates P5C and glutamate-semialdehyde (GSA), which spontaneously interconvert. P5C and GSA are also intermediates in the production of glutamate from ornithine and α-ketoglutarate catalyzed by ornithine δ-aminotransferase (OAT). ProDH and P5CDH form a fused bifunctional PutA enzyme in Gram-negative bacteria and are associated in a bifunctional substrate channelling complex in Thermus thermophilus, but the physical proximity of ProDH and P5CDH in eukaryotes has not been described. Here we report evidence of physical proximity and interactions between Arabidopsis ProDH, P5CDH and OAT in the mitochondria of plants during dark-induced leaf senescence when all three enzymes are expressed. Pairwise interactions and localization of the three enzymes were investigated using bimolecular fluorescence complementation (BiFC) with confocal microscopy in tobacco and sub-mitochondrial fractionation in Arabidopsis. Evidence for a complex composed of ProDH, P5CDH, and OAT was revealed by co-migration of the proteins in native conditions upon gel electrophoresis. Co-immunoprecipitation coupled with mass spectrometry analysis confirmed the presence of the P5C metabolism complex in Arabidopsis. Pull-down assays further demonstrated a direct interaction between ProDH1 and P5CDH. P5C metabolism complexes may channel P5C among the constituent enzymes and directly provide electrons to the respiratory electron chain via ProDH.
Keywords
- Arabidopsis thaliana, mitochondria, ornithine δ-aminotransferase, proline catabolism, proline dehydrogenase, protein complex, pyrroline-5-carboxylate dehydrogenase
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Physiology
- Agricultural and Biological Sciences(all)
- Plant Science
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In: Journal of experimental botany, Vol. 75, No. 3, 02.02.2024, p. 917-934.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Pyrroline-5-carboxylate metabolism protein complex detected in Arabidopsis thaliana leaf mitochondria
AU - Zheng, Yao
AU - Cabassa-Hourton, Cécile
AU - Eubel, Holger
AU - Chevreux, Guillaume
AU - Lignieres, Laurent
AU - Crilat, Emilie
AU - Braun, Hans-Peter
AU - Lebreton, Sandrine
AU - Savouré, Arnould
N1 - Funding Information: This work was supported by the Hubert Curien PROCOPE program between Germany and France (grant no. 46711VJ) funded by the Ministry of Foreign Affairs and by Sorbonne University. This work was also supported by a grant from the China Scholarship Council to YZ. We thank Région Ile de France for financial support of the ProteoSeine@IJM platform.
PY - 2024/2/2
Y1 - 2024/2/2
N2 - Proline dehydrogenase (ProDH) and pyrroline-5-carboxylate (P5C) dehydrogenase (P5CDH) catalyze the oxidation of proline into glutamate via the intermediates P5C and glutamate-semialdehyde (GSA), which spontaneously interconvert. P5C and GSA are also intermediates in the production of glutamate from ornithine and α-ketoglutarate catalyzed by ornithine δ-aminotransferase (OAT). ProDH and P5CDH form a fused bifunctional PutA enzyme in Gram-negative bacteria and are associated in a bifunctional substrate channelling complex in Thermus thermophilus, but the physical proximity of ProDH and P5CDH in eukaryotes has not been described. Here we report evidence of physical proximity and interactions between Arabidopsis ProDH, P5CDH and OAT in the mitochondria of plants during dark-induced leaf senescence when all three enzymes are expressed. Pairwise interactions and localization of the three enzymes were investigated using bimolecular fluorescence complementation (BiFC) with confocal microscopy in tobacco and sub-mitochondrial fractionation in Arabidopsis. Evidence for a complex composed of ProDH, P5CDH, and OAT was revealed by co-migration of the proteins in native conditions upon gel electrophoresis. Co-immunoprecipitation coupled with mass spectrometry analysis confirmed the presence of the P5C metabolism complex in Arabidopsis. Pull-down assays further demonstrated a direct interaction between ProDH1 and P5CDH. P5C metabolism complexes may channel P5C among the constituent enzymes and directly provide electrons to the respiratory electron chain via ProDH.
AB - Proline dehydrogenase (ProDH) and pyrroline-5-carboxylate (P5C) dehydrogenase (P5CDH) catalyze the oxidation of proline into glutamate via the intermediates P5C and glutamate-semialdehyde (GSA), which spontaneously interconvert. P5C and GSA are also intermediates in the production of glutamate from ornithine and α-ketoglutarate catalyzed by ornithine δ-aminotransferase (OAT). ProDH and P5CDH form a fused bifunctional PutA enzyme in Gram-negative bacteria and are associated in a bifunctional substrate channelling complex in Thermus thermophilus, but the physical proximity of ProDH and P5CDH in eukaryotes has not been described. Here we report evidence of physical proximity and interactions between Arabidopsis ProDH, P5CDH and OAT in the mitochondria of plants during dark-induced leaf senescence when all three enzymes are expressed. Pairwise interactions and localization of the three enzymes were investigated using bimolecular fluorescence complementation (BiFC) with confocal microscopy in tobacco and sub-mitochondrial fractionation in Arabidopsis. Evidence for a complex composed of ProDH, P5CDH, and OAT was revealed by co-migration of the proteins in native conditions upon gel electrophoresis. Co-immunoprecipitation coupled with mass spectrometry analysis confirmed the presence of the P5C metabolism complex in Arabidopsis. Pull-down assays further demonstrated a direct interaction between ProDH1 and P5CDH. P5C metabolism complexes may channel P5C among the constituent enzymes and directly provide electrons to the respiratory electron chain via ProDH.
KW - Arabidopsis thaliana
KW - mitochondria
KW - ornithine δ-aminotransferase
KW - proline catabolism
KW - proline dehydrogenase
KW - protein complex
KW - pyrroline-5-carboxylate dehydrogenase
UR - http://www.scopus.com/inward/record.url?scp=85184486048&partnerID=8YFLogxK
U2 - 10.1093/jxb/erad406
DO - 10.1093/jxb/erad406
M3 - Article
C2 - 37843921
VL - 75
SP - 917
EP - 934
JO - Journal of experimental botany
JF - Journal of experimental botany
SN - 0022-0957
IS - 3
ER -