Identification of protein–protein interactions of isoflavonoid biosynthetic enzymes with 2-hydroxyisoflavanone synthase in soybean (Glycine max (L.) Merr.)
| Autor: | Tomoyoshi Akashi, Toru Nakayama, Toshiyuki Waki, Seiji Takahashi, Toshio Aoki, Konstantin Denessiouk, Dong Chan Yoo, Ryo Mameda, Naoto Fujino, Reiko Motohashi, Shin-ichi Ayabe, Satoshi Yamashita |
|---|---|
| Rok vydání: | 2016 |
| Předmět: |
0106 biological sciences
0301 basic medicine Chalcone synthase Biophysics 01 natural sciences Biochemistry Isozyme Protein–protein interaction 03 medical and health sciences Bimolecular fluorescence complementation chemistry.chemical_compound Cytochrome P-450 Enzyme System Biosynthesis Isoflavonoid Protein Interaction Mapping Intramolecular Lyases Molecular Biology Flavonoids chemistry.chemical_classification biology fungi food and beverages Cell Biology Recombinant Proteins 030104 developmental biology Enzyme Flavonoid biosynthesis chemistry biology.protein Soybeans Acyltransferases 010606 plant biology & botany |
| Zdroj: | Biochemical and Biophysical Research Communications. 469:546-551 |
| ISSN: | 0006-291X |
| DOI: | 10.1016/j.bbrc.2015.12.038 |
| Popis: | Metabolic enzymes, including those involved in flavonoid biosynthesis, are proposed to form weakly bound, ordered protein complexes, called "metabolons". Some hypothetical models of flavonoid biosynthetic metabolons have been proposed, in which metabolic enzymes are believed to anchor to the cytoplasmic surface of the endoplasmic reticulum (ER) via ER-bound cytochrome P450 isozymes (P450s). However, no convincing evidence for the interaction of flavonoid biosynthetic enzymes with P450s has been reported previously. Here, we analyzed binary protein-protein interactions of 2-hydroxyisoflavanone synthase 1 (GmIFS1), a P450 (CYP93C), with cytoplasmic enzymes involved in isoflavone biosynthesis in soybean. We identified binary interactions between GmIFS1 and chalcone synthase 1 (GmCHS1) and between GmIFS1 and chalcone isomerases (GmCHIs) by using a split-ubiquitin membrane yeast two-hybrid system. These binary interactions were confirmed in planta by means of bimolecular fluorescence complementation (BiFC) using tobacco leaf cells. In these BiFC analyses, fluorescence signals that arose from the interaction of these cytoplasmic enzymes with GmIFS1 generated sharp, network-like intracellular patterns, which was very similar to the ER-localized fluorescence patterns of GmIFS1 labeled with a fluorescent protein. These observations provide strong evidence that, in planta, interaction of GmCHS1 and GmCHIs with GmIFS1 takes place on ER on which GmIFS1 is located, and also provide important clues to understand how enzymes and proteins form metabolons to establish efficient metabolic flux of (iso)flavonoid biosynthesis. |
| Databáze: | OpenAIRE |
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