Abstract
Plant secondary metabolism has been a focus of research in recent years due to its significant roles in plant defense and in human medicine and nutrition. A protein engineering strategy was designed to more effectively manipulate plant secondary metabolite (isoflavonoid) biosynthesis. A bifunctional isoflavone synthase/chalcone isomerase (IFS/CHI) enzyme was constructed by in-frame gene fusion, and expressed in yeast and tobacco. The fusion protein was targeted to the endoplasmic reticulum (ER) membrane and the individual enzymatic functions of its component fragments were retained when assayed in yeast. Petals and young leaves of IFS/CHI transgenic tobacco plants produced higher levels of the isoflavone genistein and genistein glycosides as a ratio of total flavonoids produced than did plants transformed with IFS alone. Thus, through a combined molecular modeling, in vitro protein engineering and in planta metabolic engineering approach, it was possible to increase the potential for accumulation of isoflavonoid compounds in non-legume plants. Construction of bifunctional enzymes will simplify the transformation of plants with multiple pathway genes, and such enzymes may find broad uses for enzyme (e.g., cytochrome P450 family) and biochemical pathway engineering.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 496-507 |
| Number of pages | 12 |
| Journal | Planta |
| Volume | 224 |
| Issue number | 3 |
| DOIs | |
| State | Published - Aug 2006 |
| Externally published | Yes |
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Keywords
- Chalcone isomerase
- Cytochrome P450
- Fusion enzyme
- Isoflavone synthase
- Legume
- Metabolic engineering
ASJC Scopus subject areas
- Plant Science
Cite this
Engineering isoflavone metabolism with an artificial bifunctional enzyme. / Tian, L.; Dixon, R. A.
In: Planta, Vol. 224, No. 3, 08.2006, p. 496-507.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Engineering isoflavone metabolism with an artificial bifunctional enzyme
AU - Tian, L.
AU - Dixon, R. A.
PY - 2006/8
Y1 - 2006/8
N2 - Plant secondary metabolism has been a focus of research in recent years due to its significant roles in plant defense and in human medicine and nutrition. A protein engineering strategy was designed to more effectively manipulate plant secondary metabolite (isoflavonoid) biosynthesis. A bifunctional isoflavone synthase/chalcone isomerase (IFS/CHI) enzyme was constructed by in-frame gene fusion, and expressed in yeast and tobacco. The fusion protein was targeted to the endoplasmic reticulum (ER) membrane and the individual enzymatic functions of its component fragments were retained when assayed in yeast. Petals and young leaves of IFS/CHI transgenic tobacco plants produced higher levels of the isoflavone genistein and genistein glycosides as a ratio of total flavonoids produced than did plants transformed with IFS alone. Thus, through a combined molecular modeling, in vitro protein engineering and in planta metabolic engineering approach, it was possible to increase the potential for accumulation of isoflavonoid compounds in non-legume plants. Construction of bifunctional enzymes will simplify the transformation of plants with multiple pathway genes, and such enzymes may find broad uses for enzyme (e.g., cytochrome P450 family) and biochemical pathway engineering.
AB - Plant secondary metabolism has been a focus of research in recent years due to its significant roles in plant defense and in human medicine and nutrition. A protein engineering strategy was designed to more effectively manipulate plant secondary metabolite (isoflavonoid) biosynthesis. A bifunctional isoflavone synthase/chalcone isomerase (IFS/CHI) enzyme was constructed by in-frame gene fusion, and expressed in yeast and tobacco. The fusion protein was targeted to the endoplasmic reticulum (ER) membrane and the individual enzymatic functions of its component fragments were retained when assayed in yeast. Petals and young leaves of IFS/CHI transgenic tobacco plants produced higher levels of the isoflavone genistein and genistein glycosides as a ratio of total flavonoids produced than did plants transformed with IFS alone. Thus, through a combined molecular modeling, in vitro protein engineering and in planta metabolic engineering approach, it was possible to increase the potential for accumulation of isoflavonoid compounds in non-legume plants. Construction of bifunctional enzymes will simplify the transformation of plants with multiple pathway genes, and such enzymes may find broad uses for enzyme (e.g., cytochrome P450 family) and biochemical pathway engineering.
KW - Chalcone isomerase
KW - Cytochrome P450
KW - Fusion enzyme
KW - Isoflavone synthase
KW - Legume
KW - Metabolic engineering
UR - http://www.scopus.com/inward/record.url?scp=33746888659&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=33746888659&partnerID=8YFLogxK
U2 - 10.1007/s00425-006-0233-0
DO - 10.1007/s00425-006-0233-0
M3 - Article
VL - 224
SP - 496
EP - 507
JO - Planta
JF - Planta
SN - 0032-0935
IS - 3
ER -