(Last modification:
26.10.2010)
Scots pine (Pinus sylvestris:
a few pictures from Wikipedia
More information in Wikipedia:
English:
Scots Pine ;
Deutsch:
Waldkiefer
|
Scots pine, Waldkiefer
(Pinus sylvestris) |
Young female (top right), male (middle left), and
mature cones (bottom) |
Young Cones
Bark |
Adult cone |
Crystal structure of a
stilbene synthase (STS) from Pinus sylvestris, and mechanisms of the enzyme reaction
Some
points you should know about STS
STS developed several times independently in quite
unrelated plant families
the STS Typ cyclization requires a thioesterase activity, in
contrast to CHS
the removal of the terminal carboxy group as
CO2 occurs
simultaneously with the cyclization/aromatisation (there is no carboxylated
intermediate!)
Follow the links
here or above to a few pages summarizing the new results on the mechanisms of the STS-reaction!
Key Reference
Austin,M.B.; Bowman,M.E.; Ferrer,J.-L.; Schröder,J.; Noel,J.P.: An aldol switch discovered in stilbene synthases mediates cyclization specificity of type III polyketide synthases. Chemistry & Biology 11, 1179-1194 (2004)
Stilbene synthase (STS) and chalcone synthase (CHS) each catalyze the formation of a tetraketide intermediate from a CoA-tethered phenylpropanoid starter and three molecules of malonyl-CoA, but use different cyclization mechanisms to produce distinct chemical scaffolds for a variety of plant natural products. Here we present the first STS crystal structure, and identify, by mutagenic conversion of alfalfa CHS into a functional stilbene synthase, the structural basis for the evolution of STS cyclization specificity in type III polyketide synthase (PKS) enzymes. Additional mutagenesis and enzymatic characterization confirms that electronic effects rather than steric factors balance competing cyclization specificities in CHS and STS. Finally, we discuss the problematic in vitro reconstitution of plant stilbenecarboxylate pathways, using insights from existing biomimetic polyketide cyclization studies to generate a novel mechanistic hypothesis to explain stilbenecarboxylate biosynthesis.
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or go the:
Home page of Joseph Noel; most of the work was done in his lab !!
Our other publications on the stilbene synthase (pinosylvin synthase) in Pinus sylvestris
-
Schanz, S.,
Schröder, G. and Schröder, J.:
Stilbene synthase from Scots pine (Pinus sylvestris).
FEBS Letters 313, 71-74
(1992).
Stilbene synthases are named according to their substrate
preferences. By this definition, enzymes preferring cinnamoyl-CoA are
pinosylvin synthases, and proteins with a preference for phenylpropionyl-CoA
are dihydropinosylvin synthases. We investigated the assignment of a
stilbene synthase cloned from Scots pine (Pinus sylvestris) as
dihydropinosylvin synthase and the proposal of an additional pinosylvin
synthase (1992, Plant Mol. Biol. 18, 489-503). The results show that the
previous interpretation was misled by several unexpected factors. Firstly,
we found that the substrate preference and the activity of the
plant-specific protein expressed in Escherichia coli was influenced by
bacterial factors. This was reduced by improvement of the expression system,
and the subsequent kinetic analysis revealed that cinnamoyl-CoA rather than
phenylpropionyl-CoA is the preferred substrate of the cloned stilbene
synthase. Secondly, mixing experiments showed that extracts from P.
sylvestris contain factor(s) which selectively influenced the
substrate preference, i.e. the activity was reduced with phenylpropionyl-
CoA, but not with cinnamoyl-CoA. This explained the apparent differences
between plant extracts and the cloned enzyme expressed in E. coli. Taken
together, the results indicate that the cloned enzyme is a pinosylvin
synthase, and there is no evidence for a second stilbene synthase. This
study cautions that factors in the natural and in new hosts may complicate
the functional identification of cloned sequences.
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-
Fliegmann,
J., Schröder, G., Schanz, S., Britsch, L. and Schröder, J.: Molecular analysis
of chalcone and dihydropinosylvin synthase from Scots pine (Pinus sylvestris), and differential regulation of these and related enzyme
activities in stressed plants. Plant Molecular Biology 18, 489-503 (1992).
Chalcone synthase (CHS) and stilbene synthase (STS) are closely related
polyketide synthases which are key enzymes in the biosynthesis of flavonoids and
stilbenes. Scots pine (Pinus sylvestris) is an interesting plant
for a direct comparison of the enzymes. It not only contains the usual
flavonoids, but also an unusual chalcone derivative (pinocembrin), and it
synthesizes stilbenes of the pinosylvin type. We analysed a CHS and a STS by
molecular cloning and functional expression in Escherichia coli.
The CHS was active not only with 4-coumaroyl-CoA (to naringenin chalcone), but
also with cinnamoyl-CoA (leading to pinocembrin). The STS was identified as
dihydropinosylvin synthase, because it preferred dihydrocinnamoyl-CoA to
cinnamoyl-CoA. The protein deviated in 47 positions from the CHS consensus. It
had 73.2% identity with the CHS from P. sylvestris and only 65.3%
with a STS from peanut (Arachis hypogaea). We also investigated
the regulation of both enzyme types in P. sylvestris plantlets
exposed to stress. CHS was present in non-stressed plantlets, and induction led
to a transient increase with a peak after 16 h. STS1 type activities were
regulated differently and were absent in non-stressed plantlets. Increases were
observed after a lag period of at least 6 h, and highest activities were
obtained after 30 h. The analysis of the reactions in the plant extracts and the
substrate specificity of the cloned STS suggest that the plants contain at least
two different types of STS: the cloned dihydropinosylvin synthase and a
pinosylvin synthase which preferentially utilizes cinnamoyl-CoA as substrate.
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