Indole alkaloids biosynthetic pathways

Regulation of the Terpenoid Indole Alkaloid Biosynthetic Pathway 111... 

DE LUCA, V., BALSEVICH, J., TYLER, R.T., KURZ, W.G.W, Characterization of a novel V-methyltransferase (NMT) from Catharanthus roseus plants. Detection of NMT and other enzymes of the indole alkaloid biosynthetic pathway in different cell suspension culture systems. Plant Cell Rep., 1987,6,458-461. 

Bioproduction Catharanthus roseus in vitro cultures terpenoid indole alkaloids biosynthetic pathways... 

Figure 8,2 Terpenoid indole alkaloid biosynthetic pathway. AS anthranilate synthase TDC tryptophan decarboxylase ...

MENKE, F.L., PARCHMANN, S., MUELLER, M.J., KUNE, J.W., MEMELINK, J., Involvement of the octadecanoid pathway and protein phosphoiylation in fungal elicitor-induced expression of terpenoid indole alkaloid biosynthetic genes in Catharanthus roseus. Plant Physiol., 1999,119, 1289-1296. 

Fig. 2. Alkaloid biosynthetic pathways are associated with a diverse variety of cell types. The tissue-specific localization (shaded) of enzymes and/or gene transcripts are depicted for the biosynthesis of tropane alkaloids in Atropa belladonna and Hyoscyamus niger roots (A), monoterpenoid indole alkaloids in Catharanthus roseus leaves (B), pyrrolizidine alkaloids in Senecio vernalis roots (C), pyrrolizidine alkaloids in Eupatorium cannabinum roots (D), benzyl-isoquinoline alkaloids in Papaver somniferum vascular bundles (E), and protoberberine alkaloids in Thalictrwn flamtm roots (F).

Terpenoid indole alkaloid biosynthetic enzymes are associated with at least three different cell types in C. roseus TDC and STR are localized to the epidermis of aerial organs and the apical meristem of roots, D4H and DAT are restricted to the laticifers and idio-blasts of leaves and stems, and GlOH is found in internal parenchyma of aerial organs (St-Pierre et al. 1999 Buriat et al. 2004) thus, vindoline pathway intermediates must be translocated between cell types. Moreover, enzymes involved in terpenoid indole alkaloid biosynthesis in C. roseus are also localized to at least five subcellular compartments TDC, D4H and DAT are in the cytosol, STR and the peroxidase that couples catharanthine to vinblastine are localized to the vacuole indicating transport of tryptamine across the tono-plast, SGD is a soluble enzyme associated with the cytoplasmic face of the endoplasmic reticulum, the P450-dependent monooxygenases are integral endomembrane proteins, and the N-methyltransferase involved in vindoline biosynthesis is localized to thylakoid membranes (De Luca and St-Pierre 2000). 

The in vivo transformation of [6-14C]strictosidine (19) to gelsemine in Gelsemium sempervirens was claimed with an incorporation of 0.47% (33). This provides another experimental support to the proposal that strictosidine appears to be the original precursor in the biosynthesis of monoterpenoid indole alkaloids, although the detailed pathway of this biosynthetic process still remains obscure. 

Salutaridinol 7-0-acetyltransferase catalyzes the conversion of the phenanthrene alkaloid salutaridinol to salutaridinol-7-Oacetate, the immediate precursor of thebaine along the morphine biosynthetic pathway in P. somniferum (Fig. 10.7).26 Acetyl CoA-dependent acetyltransferases have an important role in plant alkaloid metabolism. They are involved in the synthesis of monoterpenoid indole alkaloids in medicinal plant species such as Rauwolfia serpentina. In this plant, the enzyme vinorine synthase transfers an acetyl group from acetyl CoA to 16-epi-vellosimine to form vinorine. This acetyl transfer is accompanied by a concomitant skeletal rearrangement from the sarpagan- to the ajmalan-type (reviewed in2). An acetyl CoA-dependent acetyltransferase also participates in vindoline biosynthesis in Catharanthus roseus, the source of the chemotherapeutic dimeric indole alkaloid vinblastine (reviewed in2). Acetyl CoA deacetylvindoline 4-O-acetyltransferase catalyzes the last step in vindoline biosynthesis. A cDNA encoding acetyl CoA deacetylvindoline 4-0-acetyltransferase was recently successfully isolated.27... 

This overall biosynthetic scheme is summarized in Scheme 7, and it is well to remember that C. roseus is almost alone as a plant in which this whole pathway can be viewed in its entirety, for most indole alkaloid-containing plants produce only one or two of the major alkaloid classes, and not all four. In addition, C. roseus is without doubt the most economically important of the indole alkaloid-containing plants, and thus studies were, and continue to be, driven by the goal of increasing the availability of the commercially significant alkaloids ajmalicine (39), vinblastine (1), and vincristine (2). 

This review of the biosynthesis of the bisindole alkaloids of C. roseus is organized along a developing biosynthetic pathway, as far as possible, and relies on the notion that the most sophisticated studies are those utilizing the purified enzyme systems. Biosynthetic studies on the other monoterpene indole alkaloids are not reviewed here. 

The glucosidases involved in the biosynthetic pathway have been studied in detail by Hemscheidt and Zenk 203), and two of the isolated enzymes were specific for the hydrolysis of strictosidine (33) and were purified 120-fold. The enzyme was isolated from a number of indole alkaloid producing plants, including C. roseus, C.pusilus, and C. trichophyllus. The pH optimum was 6.5, and the values were 0.2 mM for enzyme I and 0.1 mM for enzyme II. Molecular weights were estimated at 230,000 for enzyme I and 450,000 for enzyme II. Unlike the case of the enzyme system of Scott et al. 198,199), tryptamine did not activate either of the two enzymes. The enzymes were highly substrate specific vincoside (85)... 

Figure 25-12 Structures and some biosynthetic pathways for some hormones, indole alkaloids, and other metabolites of tryptophan.

In summary, strictosidine (17) is likely to be the unique precursor of angustine bases (15) on the one hand and of alkaloids 7 to 13 on the other. The alternative pathway which produces cyclization to strictosidine lactam, namely, aromatiza-tion of the C ring, should most probably occur at an early stage of the biosynthetic route to the indole alkaloids of Pauridiantha. 

Within the natural products field, the investigation of complete biosynthetic pathways at the enzyme level has been especially successful for plant alkaloids of the monoterpenoid indole alkaloid family generated from the monoterpene gluco-side secologanin and decarboxylation product of tryptophan, tryptamine [3-5]. The most comprehensive enzymatic data are now available for the alkaloids ajmalicine (raubasine) from Catharanthus roseus, and for ajmaline from Indian Rauvolfia serpentina [6] the latter alkaloid with a six-membered ring system bearing nine chiral carbon atoms. Entymatic data exsist also for vindoline, the vincaleucoblastin (VLB) precursor for which some studies on enzymatic coupling of vindoline and catharanthine exist in order to get the so-called dimeric Catharanthus indole-alkaloids VLB or vincristine [7-9] with pronounced anti-cancer activity [10, 11]. 

The families Catenicellidae and Flustridae produce alkaloids derived from tryptophan. The majority of these contain bromine at carbon 6 of the indole ring, although some are more extensively brominated. The convolutamydines (brominated tryptophan derivates isolated from the genus Amathia in the order Ctenostomata)128 are similarly brominated at carbon 6, suggesting a common biosynthetic pathway or bacterial symbiont. 

Figure 7.3 Biosynthetic pathway for terpenoid indole alkaloids showing the location of enzymes for which the corresponding cDNAs have been isolated.

A large amount of structural information is available that describes the variety of indole alkaloids produced in plants. This has recently been followed by significant increases in our knowledge of the biosynthetic pathways that lead to their production and of the genes involved. Several reviews have appeared recently that describe the chemistry, biochemistry, cell and molecular biology of alkaloid biosynthesis.1 3 This chapter will selectively review recently characterized genes that appear to be responsible for the diversity and complexity of monoterpenoid indole alkaloids produced by plants. A particular focus will be on the reactions leading to the biosynthesis of vindoline (Fig. 8.1) in Catharanthus roseus. 

Some of the most interesting applications of organic structural theory to the elucidation of biosynthetic pathways were stimulated by efforts to formulate mechanisms for the biosynthesis of alkaloids. Conversely, consideration of implied biogenetic relations have occasionally helped structural determination. An important aspect of theories concerning alkaloid biosynthesis has been the assumed role of the aromatic amino acids in their formation. Only limited experimental evidence is available in this area. The incorporation of tyrosine- 8-C into morphine has been shown to be in accordance with a theory for its formation from 3,4-dihydroxyphenyl-alanine plus 3,4-dihydroxyphenylacetaldehyde. A stimulating theory of the biosynthesis of indole alkaloids, based on a condensation between trypt-amine and a rearrangement product of prephenic acid, has recently been published. The unique stereochemistry of C15 of these alkaloids had an important part in the formulation of the theory. Experimental proof of this theory would be valuable for several areas of alkaloid chemistry and biosynthesis. 

The terpenoid indole alkaloids have a variety of chemical structures and a wealth of biologic activities (Fig. 2a) (59, 60). Terpenoid indole alkaloids are used as anticancer, antimalarial, and antiarrhythmic agents. Although many biosynthetic genes from this pathway remain unidentified, recent studies have correlated terpenoid indole alkaloid production with the transcript profiles of Catharanthus roseus cell cultures (61). 

The biosynthetic pathway for ajmaline in R. serpentina is one of the best-characterized terpenoid indole alkaloid pathways. Much of this progress has been detailed in a recent extensive review (78). Like all other terpenoid indole alkaloids, ajmaline, an antiarrhythmic drug with potent sodium channel-blocking properties (79), is derived from deglycosylated strictosidine (Fig. 2c). 

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