Chemistry Reference
In-Depth Information
results after strictosidine deglycosylation, to tetrahydroalstonine
in vitro
(95).
A second
C. roseus
cell line contains an additional reductase that produces
ajmalicine. Labeling studies performed with crude
C. roseus
cell extracts in the
presence of D
2
O or NADPD support a mechanism in which the reductase acts
on the iminium form of cathenamine (96).
9.3.4 Vindoline
Vindoline, an aspidosperma-type alkaloid produced by
C. roseus
, is a key precur-
sor for vinblastine, an anticancer drug that is the most important pharmaceutical
product of
C. roseus
. Vindoline, like ajmalicine and ajmaline, is produced from
deglycosylated strictosidine. Deglycosylated strictosidine is converted to taber-
sonine through a series of biochemical steps for which no enzymatic information
exists. More details are known about the six steps that catalyze the elaboration
of tabersonine to vindoline (Fig. 9.2e) (97).
Tabersonine-16-hydroxylase, a cytochrome P450, hydroxylates tabersonine
to 16-hydroxy-tabsersonine in the first step of this sequence and has been
cloned (98, 99). The newly formed hydroxyl group is methylated by a SAM-
dependent O-methyl transferase to yield 16-methoxy-tabersonine; this enzyme
(16-hydroxytabersonine-16-O-methyltransferase) has been purified but not
cloned (100). In the next step, hydration of a double bond by an uncharacterized
enzyme produces 16-methoxy-2,3-dihydro-3-hydroxytabersonine. Transfer of
a methyl group to the indole nitrogen by an N-methyl transferase yields
desacetoxyvindoline. This methyl transferase activity has been detected only in
differentiated plants, not in plant cell cultures (101). The resulting intermediate,
deacteylvindoline, is produced by the oxoglutatarate-dependent dioxygenase
enzyme desacetylvindoline 4-hydroxylase. This enzyme has been cloned and
also is absent from plant cell cultures (102). In the last step, desacteylvindoline
is acetylated by desacteylvindoline O-acetyl transferase. This enzyme, also
absent from nondifferentiated plant material, has been cloned successfully (103).
As in morphine biosynthesis, the knowledge of the enzyme sequences allows
a more detailed understanding of the localization of the enzymes (104). Stric-
tosidine synthase (Fig. 9.2b) seems to be localized to the vacuole (105), and
strictosidine glucosidase is believed to be associated with the membrane of
the endoplasmic reticulum (73, 106). Tabersonine-16-hydroxylase is associated
with the endoplasmic reticulum membrane (98); N-methyl transferase activ-
ity is believed to be associated with the thykaloid, a structure located within
the chloroplast (101, 107); and vindoline-4-hydroxylase and desacetylvindoline
O-acetyltransferase are believed to be localized to the cytosol (Fig. 9.2e) (107,
108). Overall, extensive subcellular trafficking of biosynthetic intermediates is
required for vindoline biosynthesis.
Aside from subcellular compartmentalization, specific cell types are required
for the biosynthesis of some terpenoid alkaloids. Several enzymes involved
in the early stages of secologanin biosynthesis seem to be localized to the
phloem parenchyma, as evidenced by immunocytochemistry and
in situ
RNA
