Biosynthetic terpenoid indole alkaloid

Geerlings A, Martinez-Lozano Ibanez M, Memelink J, van der Heijden R, Verpoorte R (2000) The strictosidine 6-D-glucosidase gene from Catharanthus roseus is regulated coordinately with other terpenoid-indole alkaloid biosynthetic genes and the encoded enzyme is located in the endoplasmic reticulum. J Biol Chem 275 3051-3056... 

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

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

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. 

When plants undergo various stresses, certain secondary metabolites, including defense compounds, accumulate. Several secondary metabolites such as terpenoid indole alkaloids, indole glucosinolate, nicotine alkaloids, and polyamines are known to accumulate through the induction of biosynthetic genes by jasmonates.898-900 MeJA also induces genes involved in the formation of tryptophan derivatives, terpenoid indole alkaloids.901 These compounds are known to be involved in defense response to pathogen attack as phytoalexins. 

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). 

Transcription factors that upregulate strictosidine synthase (132), as well as a transcription factor that coordinately upregu-lates expression of several terpenoid indole alkaloid biosynthetic genes, have been found (133). Several zinc finger proteins that act as transcriptional repressors to tryptophan decarboxylase and strictosidine synthase also have been identified (134). Manipulation of these transcription factors may allow tight control of the regulation of terpenoid indole alkaloid production. Interestingly, expression of a transcription factor from Arabidopsis thaliana in C. roseus cell cultures results in an increase in alkaloid production (135). 

The biosynthetic pathway which is indicated by the above results is illustrated in Scheme 11 norlaudanosoline (70) had previously been shown to be a morphine (71) precursor. The implication of a keto-acid rather than an aldehyde in benzyliso-quinoline formation accords with observations on the biosynthesis of simpler isoquinolines and stands in contrast with the utilization of an aldehyde, sec-ologanin, in a similar reaction in the biosynthesis of terpenoid indole alkaloids. ... 

The C-3 epimeric lactams [as (154)] were proposed as potential intermediates after vincoside/isovincoside, and in support the lactam (154), synthesized from radioactive isovincoside (153), was found to be an efficient precursor for camptothecin (155) the epimeric lactam formed from radioactive vincoside (142) was only incorporated to an insignificant extent. These results stand in contrast to those for the terpenoid indole alkaloids where vincoside but not isovincoside has been found to be a biosynthetic intermediate. 

After their discovery, the Vinca alkaloids became the first natural anticancer agents to be clinically used, and they are still an indispensable part of most curative regimens used in cancer chemotherapy nowadays. On the other hand, the plant producing these alkaloids, C. roseus, has become one of the most extensively studied medicinal plants. The levels of vincristine and vinblastine in the plant revealed to be extremely low and, for pharmaceutical production, approximately half a ton of dry leaves is needed to obtain 1 g of vinblastine [4]. This feet stimulated intense investigation in alternative methods for the production of vinblastine and vincristine, namely chemical synthesis and plant cell cultures. However, chemical synthesis showed not to be viable due to the high number of transformations involved, and the anticancer alkaloids were never detected in cell cultures, which express alkaloid metabolism very poorly [5, 6]. The biosynthetic pathway of terpenoid indole alkaloids in C. roseus has also been intensively studied with the objective of developing a manipulation strategy to improve the levels of the anticancer alkaloids in the leaves of the plant [5, 7-10]. 

Fig. (3). Compartmentalization of the biosynthetic pathway of terpenoid indole alkaloids in plant cells. G10H geraniol 16-hydroxylase SLS secologanin synthase TDC tryptophan decarboxylase STR strictosidine synthase SGD strictosidine P-D-glucosidade T16H tabersonine 16-hydroxylase OMT S-adenosyl - L-methionine 16-hydroxytabereonine - 16-O-methyltransferase NMT S-adenosyl - /.-methionine 16-methoxy - 2,3-dihydro-3-hydroxytabersonine - A -methyltransferase D4H desacetoxy vindoline 4-hydroxylase DAT acetylcoenzyme A 4-O-deacetylvindoline 4-O-aeetyltransferase PRX peroxidase.

Plants possess an incredibly diverse biosynthetic capacity leading to the production of a myriad of compounds that, although not having an apparent function for fundamental life processes (growth, development and reproduction), seem to have vital roles as mediators of ecological interactions, being very important for the survival of plants. This chemical wealth is the basis of the use of plants in medicine, and is still largely unexplored. One example of application of the so called plant secondary metabolites are the terpenoid indole alkaloids of Catharanthus rose us, used in cancer therapy, and known as the Vinca alkaloids. 

Quite recently Zenk and co-workers have shown that strictosidine is also a biological precursor for terpenoid indole alkaloids of 3fl configuration. The biosynthetic conversion proceeds with loss of hydrogen at C-3, while it is retained in the formation of 3 a, Corynanthe alkaloids (107). 

Terpenoid Indole Alkaloids.—Monoterpenes have been the subject of a comprehensive review which includes these alkaloids. Indole alkaloids have been classified according to their biogenesis and another review discusses the application of tritium labelling in biosynthetic studies on these alkaloids as well as several others. 

The fact that the various steps of the biosynthetic pathway of the terpenoid indole alkaloids occur in different cell compartments implies that transport of intermediates and products is involved. Little research has been done on this transport phenomenon, except for the transport to the final storage site of the alkaloids. Deus-Neumann and Zenk (205) postulated an active and selective transport system for Catharanthus alkaloids into the... 

G. Pasquali, Regulation of the terpenoid indole alkaloid biosynthetic gene strictosidine synthase from Catharanthus roseus. Ph.D. Thesis, Leiden University, 1994. 

Elucidation of the biosynthetic pathway leading from tryptamine and secologanin, the basic precursors of terpenoid indole alkaloids, to ajmalicine, 19-epiajmalicine, and tetrahydroalstonine was reviewed by Zenk (593) and Verpoorte (594). As both ajmalicine and serpentine are produced in cell cultures, most research has involved improving yields to commercially interesting levels. We here discuss various approaches to meet this goal. 

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 ...

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). 

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