Tropane alkaloid biosynthesis pathway 



Overview of the biosynthesis:

The main precursor of the bicyclic alkamine part is L-ornithine, converted to a diamine, putrescine, by a specific decarboxylase (OrnDC). Putrescine (which can be also obtained biogenetically from arginine) is mono-N-methylated by transferase PMT and subsequently transformed into N-methylaminobutanal by diamineoxidase DAO. Next, spontaneouscyclization-dehydration takes place, with formation of the common intermediate precursor, N-methyl- Δ1-pyrrolinium cation, from which nicotine and tropane alkaloids can be formed. This monocyclic precursor is further transformed into a corresponding 4-carbon side chain .-ketoacid intermediate by the action of two acetyl coenzyme A, (AcCoA) ester molecules. The oxobutanoic acid can cyclize to exo-carboxytropinone from which derivatives of tropine, pseudotropine or ecgonine are subsequently formed.


Fig. Biosynthetic pathway of tropane alkaloids from ornithine 
(Cordell, 1981; Leete, 1989; Robins and Walton, 1993; Humphrey and O’Hagan, 2001)

Biosynthesis of cocaine:


The additional carbon atoms required for the synthesis of cocaine are derived from acetyl-CoA, by addition of two acetyl-CoA units to the N-methyl-Δ1-pyrrolinium cation. The first addition is a Mannich-like reaction with the enolate anion from acetyl-CoA acting as a nucleophile towards the pyrrolinium cation. The second addition occurs through a Claisen condensation. This produces a racemic mixture of the 2-substituted pyrrolidine, with the retention of the thioester from the Claisen condensation. In formation of tropinone from racemic ethyl [2,3-13C2]4(N methyl- 2-pyrrolidinyl)-3-oxobutanoate there is no preference for either stereoisomer. In the biosynthesis of cocaine, however, only the (S)-enantiomer can cyclize to form the tropane ring system of cocaine. The stereoselectivity of this reaction was further investigated through study of prochiral methylene hydrogen discrimination. This is due to the extra chiral center at C-2. This process occurs through an oxidation, which regenerates the pyrrolinium cation and formation of an enolate anion, and an intramolecular Mannich reaction. The tropane ring system undergoes hydrolysis, SAM-dependent methylation, and reduction via NADPH for the formation of methylecgonine. The benzoyl moiety required for the formation of the cocaine diester is synthesized from phenylalanine via cinnamic acid. Benzoyl-CoA then combines the two units to form cocaine.


Conformation through radioactive labeling experiment:

Biosynthetic pathway on which the tropane derivatives are formed has been thoroughly elucidated with the help of radioactive labeling experiments

Biosynthesis in Datura:

Tropane alkaloid biosynthesis in Datura mainly takes place in the roots (Conklin, 1976). From the site of synthesis the compounds are translocated to upper parts of the plant. Changes in alkaloid content of leaves follow the fluctuation of roots, with a delay of approximately one month (Demeyer and Dejaegere, 1989). Degradation and transformation seems to take place continuously in stems and leaves during molecule translocation to green plant parts (van de Velde et al., 1988). Within cells, the alkaloids most likely occur in the form of crystals in the vacuoles (Verzár-Petri, 1973).

Reference:


Humphrey AJ, O’Hagan D: Tropane alkaloid biosynthesis. A century old problem unresolved. Nat Prod Rep, 2001, 18, 494–502.

Oksman-Caldentey KM: Tropane and nicotine alkaloid biosynthesis – novel approach towards biotechnological production of plant derived pharmaceuticals. Curr Pharm Biotechnol, 2007, 8, 2003–2010.

Patterson S, O’Hagan D: Biosynthetic studies on tropane alkaloid hyoscyamine in Datura stramonium; hyoscyamine is stable to in vivo oxidation and is not derived from littorine via a vicinal interchange process. Phytochemistry, 2002, 61, 323–329.

Robins R, Walton N: The biosynthesis of tropane alkaloids. In: The Alkaloids. Ed. Brossi A, Academic Press, New York, 1993, 44, 115–187.