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(Last modification: 10. April 2010)
Polyketide Synthases (PKS) in Cannabis sativa
Hemp
contains flavonoids (reviewed in
Flores-Sanchez (2008b), cannabinoids (see extra:
overview of biosynthesis), stilbenoids, phenanthrenes, and spirans
(see for example Shoyama et al., 1978;
ElSohly et al., 1982; Crombie
et al., 1982; Elsohly et al., 1984;
Crombie 1986), and for a recent general review
see:
Flores-Sanchez (2008b).
One could postulate three different activities for their biosynthesis:
-
Flavonoids: The key
reaction in biosynthesis is likely to be a standard
chalcone synthase (CHS): 4-coumaroyl-CoA as substrate, three condensations
with malonyl-CoA, and ring-folding to a chalcone. More details on the CHS
reaction are found here.
-
Cannabinoids: The initial reaction in the biosynthesis of the most
dominant compounds (a short description of the other
reactions in their
biosynthesis is here) must be
the OLivetolic
Acid
Synthase (OLAS): an enzyme using hexanoyl-CoA, carrying
out three condensations, followed by a stilbene synthase (STS)-type
ring-closure, but without the loss of the terminal carboxyl group that is
typical for standard STS-reactions. See Fig. 1 for the
postulated reaction. It should be noted that compounds derived from
butyryl-CoA are also known, but except for the different chain length of the
starter the reactions should be pretty much the same.
Actually, the reaction should be grouped with the stilbenecarboxylate
synthases because these also retain the carboxyl group: Stilbene synthase (STS) type ring-folding with retention of
the terminal carboxyl group (stilbenecarboxylate synthases, STCS) are known from other plants, see in this
website: Hydrangea macrophylla,
Marchantia polymorpha,
and in the biosynthesis postulated for
anacardic acid and urushiols (more...). However, there is one important difference: In
those cases the
reaction sequence to the carboxylated end product contains a reduction step.
That is not the case in the biosynthesis of olivetolic acid; this is potentially
important because it has been argued that reduction and retention of carboxylic
group may be linked: more....
I left it here because these enzymes should be discussed together with the
bibenzyl synthases (BBS) from Cannabis that are discussed below.
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Stilbenoids, phenanthrenes, and spirans:
The structures (Fig. 2)
suggest that their biosynthesis should be initiated by
BiBenzyl
Synthases
(BBS, essentially STS-type enzymes) using
dihydro-4-coumaroyl-CoA or dihydrocaffeoyl-CoA
as starters, with three condensations followed by a STS-type ring-folding.
See Fig. 2 for the
postulated initial reactions.
Type III PKS are excellent candidates for all three
reactions. The following summarizes the evidence available for the enzyme activities and
the molecular data.
Fig. 1:
Olivetolic Acid
Proposed Type III PKS reaction in Cannabis sativa.
The polyketide synthase uses hexanoyl-CoA
as starter and performs three condensations which are followed by a
stilbenecarboxylate (STCS)-type ring-closure, i.e. with retention of the
terminal carboxyl group. The colours indicate the carbon atoms introduced by the
three condensation reactions.
Fig. 2:
Bibenzyls
Model for the biosynthesis of some
stilbenoids, phenanthrenes, and spirans in hemp (Cannabis sativa).
The coloured arrows and dots mark the condensation reactions and the carbon
atoms introduced by them. Dotted arrows: several reactions. Note that the order
of the reactions has not been determined; the biosynthetic relationships
suggested here are just possibilities. The scheme is a simplified, modified version of Fig. 9
in the review by Flores-Sanchez and Verpoorte (2008b).
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What do we know about
these enzymes?
A candidate enzyme activity for the enzyme in
cannabinoid biosynthesis was identified in crude extracts from
plants (Raharjo et al., 2004a).
The enzyme indeed carried out a STS-type reaction; however, the reaction product was not the expected olivetolic acid
(enzyme = OLAS, olivetolic acid
synthase),
but the decarboxylation product olivetol (enzyme =
OLS, olivetol synthase). That could not possibly be the
enzyme in cannabinoid biosynthesis because it seems quite definite that the
carboxyl group is retained until the end of the biosynthetic pathway. The
subsequent enzymes, most importantly the prenyltransferase (discussed
here), does not accept olivetol, and the same is true for the next reactions (cannabinoid
synthases, discussed
here). It seems clear that the removal of the
carboxyl group occurs non-enzymatically in vitro during storage and
processing of the drug.
The same group cloned a type III PKS from Cannabis sativa
(accession: AAL92879)
(Raharjo et al., 2004b), but the recombinant enzyme had only chalcone synthase type
activities, with both the typical CHS and valerophenone synthase (VPS)
substrates. The hoped for STCS/STS-type products (4-coumaroyl-CoA -> stilbenecarboxylate or
resveratrol; hexanoyl-CoA -> olivetolic acid or olivetol) were not detected.
A recent publication described
renewed attempts by measurements of enzyme activities in crude extracts
(Flores-Sanchez and Verpoorte, 2008a). They
found the activities of chalcone synthase (CHS, tested with 4-coumaroyl-CoA),
phlorisovalerophenone synthase
(VPS), isobutyrophenone synthase (BUS),
stilbene synthase (STS, tested with 4-coumaroyl-CoA -> product resveratrol), and olivetol synthase
(OLS, with hexanoyl-CoA). Again, OLAS activity leading to olivetolic acid
was not detected. Taking the results reported for the cloned
type III PKS
(Raharjo
et al., 2004b), and considering the substrate promiscuity of these enzymes (more...),
the data suggested:
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CHS-, VPS-, and BUS-
activities: one and the same enzyme
may well
be responsible for these activities. VPS or BUS products are not known from
Cannabis sativa, but various flavonoids. It seems likely that the
cloned enzyme is a chalcone synthase in vivo.
-
Stilbene synthase (STS)- and Olivetol
synthase (OLS) activities in vitro: the significance remains
unclear. One would tend to think that this cannot be side activities of the
postulated olivetolic acid (OLAS) enzyme, because that enzyme should not
remove the terminal carboxyl group. Stilbenes in their strict
definition are not known from C. sativa, but stilbenoids and other
products that are likely derived from an STS-type reaction, but not with
4-coumaroyl-CoA: the substrates for the initial key reaction should lack the
double bond in the side chain (see Fig. 2). If so, the
authors used the wrong substrates (4-coumaroyl-CoA, hexanoyl-CoA). This may well be important:
a pronounced substrate discrimination has
been shown for the bibenzyl synthases (BBS) from orchids (see for example
Preisig-Müller et al., 1995),
and the difference in the presence/absence of the double bond in the propanoid
side chain may lead to different
product specificities. This, for example, was also observed with the stilbenecarboxylate synthases (STCS)
from Hydrangea macrophylla which were investigated with many more
substrates (more...).
In February 2009, the databases contained two entries for type III PKS in Cannabis sativa:
the CHS (AAL92879)
(Raharjo et
al., 2004b), and an entry labelled as olivetol synthase (see
below,
OLS,
BAG14339, date: 25-MAR-2008,
Submitted 01-MAR-2004),
it was at that time cited as unpublished (this is the protein with a
preliminary crystallization, Taguchi et al., 2008). The closest relation to that
protein (about 74% identity) seems to be a CHS from hop (Humulus lupulus,
BAB47196), a close relative of Cannabis sativa. The known CHS from Cannabis sativa does not even show up as closely
related in simple Blast searches. The distant relationship to the C. sativa
CHS and other typical CHS (have a look at the
relationship tree) would indeed be expected for a non-CHS activity. The
crystallization publication in fact states in its introduction that the protein
used hexanoyl-CoA to synthesize
hexanoyl triacetic acid lactone (HTAL): this
is the product from three condensations and a pyrone ring-folding (in
contrast to chalcone or stilbene ring-folding); it corresponds to the
coumaroyltriacetic acid lactone (CTAL) synthesized from 4-coumaroyl-CoA (more...).
Such lactones have been proposed as precursors of resorcinolic acids (more...):
is it possible that HTAL is an in vitro precursor of olivetolic acid, as
discussed in Taguchi et al. (2008)?
Update
17. June 2009:
Taura et al. (2009)
The functional characterization of that
enzyme discussed above has now been
published (Taura
et al., 2009). The results with the recombinant protein showed that olivetol
was the dominant reaction product, not hexanoyl triacetic acid lactone.
Olivetolic acid was not among the products. The OLS
accepted starter CoA esters with C4 to C8 side
chains such as butyryl-, isovaleryl-, and octanoyl-CoA; however, it produced
triketide pyrones from these substrates except producing 5-propylresorcinol (divarinol)
from butyryl-CoA. No product at all was detected with aromatic CoA-esters,
including 4-coumaroyl-CoA. These specificities are somewhat more narrow than
with standard CHS or STS. Most importantly, however, these characteristics do
not fit the activities expected for olivetolic acid synthesis (no carboxylated
product was detectable). The authors also tested the activities in crude extracts
from the plants. Again, olivetolic acid synthase activity was not detected; the
product was always olivetol. It is noteworthy that the highest activities were
found in flowers and rapidly expanding leaves, that means, in the tissues
producing the cannabinoids. All of this remains rather puzzling. The authors
argue that the unexpected decarboxylation might be a result of the assay
conditions in vitro, and I would tend to agree. But what is the factor
hindering the decarboxylation in vivo? Another possible explanation would
be that this enzyme is actually a bibenzyl synthase (BBS) in vivo: If so,
4-coumaroyl-CoA again would have been the wrong substrate. Unfortunately, not
even the commercially available substrate phenylpropionyl-CoA was tested: from
all our experience with this type of enzyme, one would expect a rather high
activity for the STS-type product.
An interesting question: if this is actually an enzyme with STS-type
ring-folding: will the mechanism follow the 'aldol switch mechanism' established
for the Pinus sylvestris STS?
Click here for a discussion.
Update
10. September 2009:
Marks et al. (2009)
Two points should be noted: a)
the authors did not know about the Taura et al. (2009) publication: that
appeared in June, and the Marks et al. manuscript was accepted in the same month,
and b) the main point was not PKS activities, but a general search for genes in
cannabinoid biosynthesis.
Nevertheless, a considerable part dealt with candidate PKS
sequences. Three were identified: one (= CAN1069) was the previously described
CHS
(Raharjo
et al., 2004b), another one (=
CAN24) was the OLS described by the Taura group (Taura
et al., 2009), and there was a third, new
sequence (= CAN383). Unfortunately, the analysis of the products with
recombinant protein was less than satisfactory. In particular, the CAN24 and
CAN383 products remain obscure: they did not fit the properties of olivetol
or olivetolic acid, but beyond that there is just a speculation that they may
have been
pyrones from two condensations. The only substrates investigated were
4-coumaroyl-CoA and hexanoyl-CoA. A look at the assay conditions seems to
suggest that they were rather unusual: 0.25 mM hexanoyl-CoA and 5 mM malonyl-CoA
appear very high; in particular the malonyl-CoA concentration (about 50x higher
than in most other publications). In our experience, such high concentrations
had with most of these enzymes a number of undesirable effects on the product
formation, e.g. substrate inhibition and preferential formation of derailment
products.
In
summary:
One would postulate three PKS activities in Cannabis sativa: CHS, STS (which
in vivo most
likely functions as BBS, i.e. a bibenzyl synthase), and a second STS-type enzyme
(OLAS) producing olivetolic acid, a carboxylated product. There are at
least two problems at present identifying the correct enzymes: a)
the BBS function should not be tested with the substrates used so far
(4-coumaroyl-CoA and hexanoyl-CoA), and
b) it remains a mystery why the protein identified as having OLS
activity does not produce olivetolic acid: all the evidence from tissue-specific
expression and its intensity argue that it should be the olivetolic acid
synthase. However, the production of carboxylated products with STS-type enzymes
seems to be tricky in general: more...
Would a transgenic expression help to solve the problem? Maybe,
but it should be ensured that the postulated
CoA-esters are actually available in the transformed plants.
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Links zu STS-Typ Enzymen
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Marks, M. D., Tian, L., Wenger, J. P., Omburo, S.
N., Soto-Fuentes, W., He, J., Gang, D. R., Weiblen, G. D., Dixon, R. A.,
2009.
Identification of candidate genes affecting
D9-tetrahydrocannabinol
biosynthesis in Cannabis sativa. Journal of Experimental Botany 60,
3715-3726.
RNA isolated from the glands of a
D9-tetrahydrocannabinolic
acid (THCA)-producing strain of Cannabis sativa was used to generate
a cDNA library containing over 100 000 expressed sequence tags (ESTs).
Sequencing of over 2000 clones from the library resulted in the
identification of over 1000 unigenes. Candidate genes for almost every step
in the biochemical pathways leading from primary metabolites to THCA were
identified. Quantitative PCR analysis suggested that many of the pathway
genes are preferentially expressed in the glands. Hexanoyl-CoA, one of the
metabolites required for THCA synthesis, could be made via either de novo
fatty acids synthesis or via the breakdown of existing lipids. qPCR analysis
supported the de novo pathway. Many of the ESTs encode transcription
factors and two putative MYB genes were identified that were preferentially
expressed in glands. Given the similarity of the Cannabis MYB genes to those
in other species with known functions, these Cannabis MYBs may play roles in
regulating gland development and THCA synthesis. Three candidates for the
polyketide synthase (PKS) gene responsible for the first committed step in
the pathway to THCA were characterized in more detail. One of these was
identical to a previously reported chalcone synthase (CHS) and was found to
have CHS activity. All three could use malonyl-CoA and hexanoyl-CoA as
substrates, including the CHS, but reaction conditions were not identified
that allowed for the production of olivetolic acid (the proposed product of
the PKS activity needed for THCA synthesis). One of the PKS candidates was
highly and specifically expressed in glands (relative to whole leaves) and,
on the basis of these expression data, it is proposed to be the most likely
PKS responsible for olivetolic acid synthesis in Cannabis glands.
Interesting publication, but some points are a bit annoying:
- just do not take the structures of the CoA-esters in Fig. 5 seriously:
they are wrong (-O-SCoA instead of the correct -SCoA).
- it seems a bit strange that the same mistake appears in several other
publications on Cannabis PKS, from an independent group:
- Flores-Sanchez and Verpoorte (2008a and 2008b); Raharjo et al. (2004)
- Fig. 2: n-Hexanol (an alcohol!) is not a product of fatty acid
biosynthesis or degradation: it is the acid, not the alcohol. And of course,
the acyl-CoA synthetases use the acid as substrate, not the alcohol.
- the product identifications are less than convincing, beyond the statement
that olivetol or olivetolic acid were not among the products.
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Taura, F., Tanaka, S., Taguchi, C., Fukamizu, T.,
Tanaka, H., Shoyama, Y., Morimoto, S., 2009. Characterization of olivetol
synthase, a polyketide synthase putatively involved in cannabinoid
biosynthetic pathway.
FEBS Letters 583, 2061-2066.
Alkylresorcinol moieties of cannabinoids are derived from olivetolic acid (OLA),
a polyketide metabolite. However, the polyketide synthase (PKS) responsible
for OLA biosynthesis has not been identified. In the present study, a cDNA
encoding a novel PKS, olivetol synthase (OLS), was cloned from Cannabis
sativa. Recombinant OLS did not produce OLA, but synthesized olivetol,
the decarboxylated form of OLA, as the major reaction product.
Interestingly, it was also confirmed that the crude enzyme extracts from
flowers and rapidly expanding leaves, the cannabinoid-producing tissues of
C. sativa, also exhibited olivetol-producing activity, suggesting
that the native OLS is functionally expressed in these tissues. The
possibility that OLS could be involved in OLA biosynthesis was discussed
based on its catalytic properties and expression profile.
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Flores-Sanchez, I. J., Verpoorte, R.,
2008a. PKS activities and biosynthesis of cannabinoids and flavonoids in
Cannabis sativa L. plants. Plant and Cell Physiology 49, 1767-1782.
Polyketide
synthase (PKS) enzymatic activities were analyzed in crude protein extracts
from cannabis plant tissues. Chalcone synthase (CHS, EC 2.3.1.74), stilbene
synthase (STS, EC 2.3.1.95), phlorisovalerophenone synthase (VPS, EC
2.3.1.156), isobutyrophenone synthase (BUS) and olivetol synthase activities
were detected during the development and growth of glandular trichomes on
bracts. Cannabinoid biosynthesis and accumulation take place in these
glandular trichomes. In the biosynthesis of the first precursor of
cannabinoids, olivetolic acid, a PKS could be involved; however, no activity
for an olivetolic acid-forming PKS was detected. Content analyses of
cannabinoids and flavonoids, two secondary metabolites present in this
plant, from plant tissues revealed differences in their distribution,
suggesting a diverse regulatory control for these biosynthetic fluxes in the
plant.
(Note: the structures of the CoA-esters in Fig. 1 are not correct).
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Flores-Sanchez, I. J., Verpoorte, R.,
2008b. Secondary metabolism in cannabis. Phytochemistry Reviews 7, 615-639.
Cannabis
sativa L. is an annual dioecious plant from Central Asia. Cannabinoids,
flavonoids, stilbenoids, terpenoids, alkaloids and lignans are some of the
secondary metabolites present in C. sativa. Earlier reviews were
focused on isolation and identification of more than 480 chemical compounds;
this review deals with the biosynthesis of the secondary metabolites present
in this plant. Cannabinoid biosynthesis and some closely related pathways
that involve the same precursors are discussed.
(Note: the structures of the CoA-esters in some figures are not correct).
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Go to Fig. 2
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Taguchi, C., Taura, F., Tamada, T., Shoyama, Y.,
Shoyama, Y., Tanaka, H., Kuroki, R., Morimoto, S., 2008. Crystallization and
preliminary X-ray diffraction studies of polyketide synthase-1 (PKS-1) from
Cannabis sativa. Acta Crystallographica Section F: Structural Biology
and Crystallization Communications 64, 217-220.
Polyketide
synthase-1 (PKS-1) is a novel type III polyketide synthase that catalyzes
the biosynthesis of hexanoyl triacetic acid lactone in Cannabis sativa
(Mexican strain). PKS-1 was overproduced in Escherichia coli,
purified and finally crystallized in two different space groups. The crystal
obtained in 0.1 M HEPES buffer pH 7.5 containing 0.2 M calcium acetate and
20%(w/v) polyethylene glycol 3350 diffracted to 1.65 A resolution and
belonged to space group P1, with unit-cell parameters a = 54.3, b = 59.3, c
= 62.6 A, alpha = 69, beta = 81, gamma = 80 degrees. Another crystal
obtained in 0.1 M HEPES buffer pH 7.5 containing 0.2 M sodium chloride and
20%(w/v) polyethylene glycol 3350 diffracted to 1.55 A resolution and
belonged to space group P2(1)2(1)2(1), with unit-cell parameters a = 54.3, b
= 110, c = 130 A. These data will enable us to determine the crystal
structure of PKS-1.
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Raharjo, T. J., Chang, W.
T., Choi, Y. H., Peltenburg-Looman, A. M. G., Verpoorte, R., 2004a.
Olivetol as product of a polyketide synthase in Cannabis sativa L.
Plant Science 166, 381-385.
A
polyketide synthase (PKS) was suggested to catalyze the first step of
cannabinoid biosynthesis, leading to olivetolic acid. An activity of a
PKS was detected in the protein extract of Cannabis sativa
flowering top. The enzyme converts one molecule of n-hexanoyl-CoA and
three molecules of malonyl-CoA to olivetol. The product was identified
by its UV-spectrum, mass spectrometry analysis and comparison with
reference compound. The activity of the enzyme was also found in the
upper leaves, but the activity occurring there is lesser than in the one
occurring in the flowers. The activity of chalcone synthase (CHS),
another PKS enzyme, was also found in the protein extract.
-> if you read the paper: ignore that the structure of the CoA-esters in
Fig. 1 is wrong.
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Raharjo, T. J., Chang, W. T.,
Verberne, M. C., Peltenburg-Looman, A. M. G., Linthorst, H. J. M.,
Verpoorte, R., 2004b. Cloning and over-expression of a cDNA encoding a
polyketide synthase from Cannabis sativa. Plant Physiology and
Biochemistry 42, 291-297.
A polyketide synthase has been suggested to play an important role in
cannabinoid biosynthesis in Cannabis sativa L. This enzyme
catalyzes the biosynthesis of olivetolic acid, one of the precursors for
cannabinoid biosynthesis. Using a reverse transcriptase-polymerase chain
reaction (RT-PCR) based on the DNA homology of chalcone synthase (EC
2.3.1.156) and valerophenone synthase (EC 2.3.1.156) of hop (Humulus
lupulus), a cDNA encoding a polyketide synthase in C. sativa
was identified. The coding region of the gene is 1170 bp long encoding a
389 amino acid protein of a predicted 42.7 kDa molecular mass and with a
pI of 6.04. The gene shares a high homology with a chalcone
synthase gene of H. lupulus, 85% and 94% homology on the level of
DNA and protein, respectively. Over-expression of the construct in
Escherichia coli M15 resulted in a 45 kDa protein. The protein has
chalcone synthase activity as well as valerophenone synthase activity, a
chalcone synthase-like activity. Using n-hexanoyl-CoA and
malonyl-CoA as substrates did not give olivetol or olivetolic acid as a
product.
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Raharjo, T. J., Widjaja, I., Roytrakul, S.,
Verpoorte, R., 2004c.
Comparative proteomics of Cannabis sativa plant tissues. Journal
of Biomolecular Techniques 15, 97-106.
Comparative proteomics of leaves, flowers, and glands of Cannabis
sativa have been used to identify specific tissue-expressed
proteins. These tissues have significantly different levels of
cannabinoids. Cannabinoids accumulate primarily in the glands but can
also be found in flowers and leaves. Proteins extracted from glands,
flowers, and leaves were separated using two-dimensional gel
electrophoresis. Over 800 protein spots were reproducibly resolved in
the two-dimensional gels from leaves and flowers. The patterns of the
gels were different and little correlation among the proteins could be
observed. Some proteins that were only expressed in flowers were chosen
for identification by matrix-assisted laser desorption/ionization
time-of-flight mass spectrometry and peptide mass fingerprint database
searching. Flower and gland proteomes were also compared, with the
finding that less then half of the proteins expressed in flowers were
also expressed in glands. Some selected gland protein spots were
identified: F1D9.26-unknown prot. (Arabidopsis thaliana),
phospholipase D beta 1 isoform 1a (Gossypium hirsutum), and PG1 (Hordeum
vulgare). Western blotting was employed to identify a polyketide
synthase, an enzyme believed to be involved in cannabinoid biosynthesis,
resulting in detection of a single protein.
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Preisig-Müller, R., Gnau, P., Kindl, H.,
1995.
The
inducible 9,10-dihydrophenanthrene pathway: characterization and expression
of bibenzyl synthase and S-adenosylhomocysteine hydrolase. Archives
of Biochemistry and Biophysics 317, 201-207.
Tricyclic 9,10-dihydrophenanthrenes originate from phenylpropane derivatives
by chain elongation and cyclization according to the polyacetate rule.
Bibenzyls are bicyclic intermediates, and O- methylation is a
prerequisite for their conversion into dihydrophenanthrenes. cDNA clones
encoding bibenzyl synthases and S-adenosylhomocysteine hydrolase of
the orchid Phalaenopsis sp. were isolated from a cDNA library
representing the stage of elicitor-induced plants. The deduced amino acid
sequences of two clones, pBibSy811 and pBibSy212, indicated that we obtained
two full-length sequences of bibenzyl synthases characterized by their
homology to stilbene synthases previously investigated. That indeed bibenzyl
synthase cDNAs rather than a homologous stilbene synthase cDNA or chalcone
synthase cDNA have been isolated was demonstrated by expression of two
enzymatically active bibenzyl synthase proteins in Escherichia coli.
These proteins showed virtually the same selectivity towards
m-hydroxyphenylpropionyl-CoA as substrate as the enzyme isolated from orchid
plants. In young sterile Phalaenopsis plants, the formation of both
bibenzyl synthase mRNAs and S-adenosylhomocysteine hydrolase mRNAs
was increased upon elicitation more than 100-fold. The time courses of gene
expression exhibited transient profiles, reaching maximum mRNA levels 20 h
after onset of fungal infection followed by a rapid decline to 40 h.
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Crombie, L., 1986. Natural products of cannabis
and khat. Pure and Applied Chemistry 58, 693-700.
The chemistry of
two drugs of abuse is surveyed. Cannabis sativa contains two major
series of natural products: first, the cannabinoid group which includes the
psychotomimetic Li1-THC, and second, a biogenetically connected series
involving bibenzyls, spiro-compounds, dihydrophenanthrenes and flavonoids.
At an early biogenetic stage there are connections between these two series,
and late stage 'chemical crossing' is described. The E.African drug Khat (Catha
edulis) is used in Arab lands, but in contrast to Cannabis much less is
known of its pharmacology. Khat contains the stimulants cathine and
cathinone, but chemical interest centres particularly on a series of large
alkaloids the plant contains. These are based on highly hydroxylated
terpenic cores, derived from dihydroagarofuran, which are esterified with a
variety of acids, some forming macrocyclic bridges.
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ElSohly, H. N., Ma, G. E., Turner, C. E., ElSohly,
M. A., 1984. Constituents of Cannabis sativa, XXV. Isolation of two
new dihydrostilbenes from a Panamanian variant. Journal of Natural Products
47, 445-452.
Two new dihydrostilbene compounds (named cannabistilbenes I and II) were
isolated from a polar acidic fraction of a Panamanian variant of
Cannabis sativa grown in Mississippi. The structure of cannabistilbene
I was shown to be
3,4'-dihydroxy-5-methoxy-3'-(3-methylbut-2-enyl)-dihydrostilbene (1) from
spectral data which was confirmed by synthesis. There is spectral evidence
to indicate that cannabistilbene II could be represented by either structure
3 or 4.
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Crombie, L., Crombie, W. M. L., 1982. Natural
products of Thailand high (delta)1-THC-strain Cannabis. The
bibenzyl-spiran-dihydrophenanthrene group: relations with cannabinoids and
canniflavones. Journal of the Chemical Society, Perkin Transactions 1,
1455-1466.
Since non-cannabinoids may influence the pharmacological profile of
Cannabis-leaf drug, a detailed examination of the acidic fraction from leaf
extractive has been made. Twelve non-cannabinoids have been isolated
crystalline from a single high 1-THC-strain of Thailand Cannabis grown in
Nottingham under controlled conditions: nine of the compounds were not
previously known as natural products and their structures have been
determined. The extractives comprise three bibenzyls, six spirans, two
9,10-dihydrophenanthrenes, and two prenylated flavones.The bibenzyls,
spirans, and dihydrophenanthrenes may be linked together in a biogenetic
scheme in which one-electron oxidation and reductive processes play
important parts: the scheme is particularly supported by the discovery of a
new spiran, cannabispiradienone, which holds a key position and undergoes a
dienone-phenol rearrangement to give one of the new dihydrophenanthrenes.
Relations between bibenzyl, cannabinoid, and flavone pathways are briefly
considered.
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Elsohly, H. N., Turner, C. E., 1982. Constituents
of Cannabis sativa L. XXII: isolation of spiro-indan and
dihydrostilbene compounds from a Panamanian variant grown in Mississippi,
United States of America. Bulletin on Narcotics 34, 51-56.
Three spiro-compounds, namely cannabispiran, dehydrocannabispiran and
beta-cannabispiranol, and 2 dihydrostilbenes
[3-(2-(3-hydroxy-4-methoxyphenyl)ethyl)-5-methoxyphenol and canniprene] were
isolated from a polar fraction of a Panamanian variant of Cannabis sativa
L. grown in Mississippi, United States of America. The plant material was
extracted with 95% ethanol and the dried ethanol extract was then
partitioned between chloroform and water. The chloroform fraction was
fractionated between hexane and 3N sodium hydroxide solution. Acidification
of the basic fraction followed by extraction with ether afforded a polar
acidic fraction from which the above-mentioned compounds were isolated
through repeated chromatography. The structures of the above compounds were
determined by spectral means as well as by comparison with reference
samples. The isolation of two dihydrostilbenes and three spiro-indan
compounds from a single variant provides good support that the
dihydrostilbenes are the natural precursors to the spiro-indan compounds.
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Shoyama, Y., Nishioka, I., 1978. Cannabis. XIII.
Two new spiro-compounds, cannabispirol and acetyl cannabispirol. Chemical &
Pharmaceutical Bulletin (Tokyo) 26, 3641-3646.
Two new spiro-compounds, cannabispirol and acetyl cannabispirol, were
isolated along with cannabispirone and cannabispirenone from the Japanese
domestic cannabis and these structures were elucidated. The biogenetic
relationship of spiro-compounds and cannabinoids was also discussed.
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