Biosynthesis elymoclavine

Ergot Alkaloids.—4-(yy-Dimethylallyl)tryptophan (122) is the first intermediate beyond tryptophan in ergot alkaloid biosynthesis. Chanoclavine-I (127) is the first tricyclic base (cf. Vol. 10, p. 26, and ref. 2). Recently, (124 labels as shown) has been found to be a very efficient and intact precursor for elymoclavine (128).45 The high level of incorporation indicates that (123) is a probable intermediate situated between (122) and (127). The decarboxylation product (125) was not utilized for biosynthesis, so, although decarboxylation of (123) is required for the conversion of (123) into (127), either it is intimately associated with ring-closure or an imine that is related to (126) is involved. 

Experiments have been carried out with elymoclavine (73) in whole cells and protoplasts of Claviceps strain SD 58 levels of dimethylallyltryptophan synthetase, the first enzyme in ergot alkaloid biosynthesis, were measured.64 The results that were obtained provide strong evidence that there is end-product regulation of the synthesis of alkaloids in vivo and that it involves feedback inhibition end-product repression of the synthesis of enzymes appears to be of lesser importance. 

Figure 2.14 Biosynthesis of agroclavine and elymoclavine in Claviceps purpurea cultures.

Kobayashi, M. and Floss, H.G. (1987) Biosynthesis of ergot alkaloids origin of the oxygen atoms in chanoclavine 1 and elymoclavine.. Org. Chem., 52,4350-2. 

In the early stages of the biosynthesis of agroclavine (7) and elymoclavine (8) a metabolic grid appears to operate. Dimethylallyltryptamine (9) and its... 

A fascinating problem in the biosynthesis of ergot alkaloids is the mechanism by which dimethylallyltryptophan (102) is transformed through chanoclavine-I (103) to agroclavine (104) and elymoclavine (106), a reaction sequence which involves two cis-trans isomerizations in the side-chain double bond. Further evidence on the course of biosynthesis comes from feeding dimethylallyltryptophan labelled as shown in (102), to Claviceps penniseti. The alkaloids (103), (104), and (106) were isolated and the sites of labelling were determined to be as shown in Scheme 7. Thus the first double bond isomerization occurs between (102) and (103) and the second between (103) and (104). 

Dimethylallyltryptophan (139) has been identified as the first intermediate after tryptophan in ergot alkaloid biosynthesis.In experiments with a Claviceps species, clavicipitic acid (140) was identified as a major product formed from (139).(The enzyme catalysing this transformation was identified in both the supernatant and microsonal fractions oxygen is necessary for the reaction but cytochrome P-450 does not appear to be involved.) But radioactive clavicipitic acid was found to be a much less efficient precursor than (139) for elymoclavine and thus is in all probability not an intermediate in ergot alkaloid biosynthesis. 

Biosynthesis of the ergot bases involves an N-methylation step and it has been shown that the N-methyl group arises intact from methionine. It is as yet unknown, however, at which stage the methylation occurs but since iV-methyl-tryptophan and iV-methyltryptamine are not intermediates, the reaction must occur at a stage between (118) and chanoclavine-I (119). The failure of radioactive A -demethylchanoclavine-1 (or N-demethylchanoclavine-II) to label elymoclavine (123) in Claviceps cultures indicates that iV-methylation occurs before formation of the chanoclavine skeleton. 

The synthesis of dimethylallyltryptophan (132) by a crude extract of Claviceps purpurea from tryptophan and dimethylallyl pyrophosphate recorded earlier has been reported again recently. In addition to (132), the formation of (133) was observed. (The latter compound, with unspecified stereochemistry around the double bond, has also been isolated from a C. purpurea culture ). It was found further that both (132) and (133) could act as precursors for lysergic acid amides in C. paspali cultures. Both (133) and its (Z)-isomer have been found to act as precursors for elymoclavine (137) but not chanoclavine-I (138) or agro-clavine (136), which are considered to be normal intermediates in elymoclavine biosynthesis.It may be concluded, however, from the combined evidence, that elymoclavine, lysergic acid, and related compounds may normally be formed along an alternative pathway via these allylic hydroxy-compounds. 

The biosynthesis of ergot alkaloids is initiated by prenyla-tion of tryptophan with dimethylallyl pyrophosphate (DMAPP) to yield 4-dimethylallyltryptophane (DMAT) (4) (Fig. 35.2) (Floss, 1980). This alkylation step is believed to be rate limiting, and the enzyme that catalyzes this reaction, DMAT synthetase, has been purified more than 60 times (Maier and Groger, 1976 Waller and Dermer, 1981). The enzyme appears to be specific for L-tryptophan and is inhibited by agroclavine (2) and elymoclavine (5) the end products of the reaction. 

To elucidate further the mechanism of closure for ring C, Floss et al. synthesized the racemic amino acid shown in Scheme 58 [88]. Feeding experiments with the N-trideuteriomethyl labeled compound showed a 33% specific incorporation of the amino acid into elymoclavine. The high level and specificity of incorporation (no M-i-1 or M-i-2 species were detected) clearly point toward the intermediacy of the tertiary carbinol in the biosynthesis of the tetracyclic ergoline ring system. On the basis of these results. Floss et al. proposed the overall sequence shown in Scheme 53 for elaboration of the C ring system [88]. 

In the course of their studies on the biosynthesis of ergot alkaloids. Robbers and Floss isolated clavidpitic acid, a new indolic amino acid [89]. Based on the X-ray crystal data of the major isomer and the assumption that the stereocenter at C-5 retains its stereochemistry from L-tryptophan, the structure shown in Scheme 59 was assigned [90]. By performing feeding experiments on Claviceps sp. SD 58 with clavicipitic acid (biosynthetically labeled), the laboratories of Floss [90] and Anderson [91] independently determined that clavicipitic acid is not a precursor to elymoclavine. Instead, Floss argues that it arises from ... a derailment of the metabolism leading to the tetracyclic ergolines between the first and second pathway-specific steps, the isoprenylation of tryptophan and the N-methylation of 4-(y,y-dimethylallyl)-tryptophan (DMAT). Anderson and co-workers isolated an enzyme, DMAT oxidase, from Claviceps sp. which catalyzes the formation of clavicipitic acid from DMAT [92]. The enzyme, which requires... 

Agurell, S. (1966b) Biosynthesis of ergot alkaloids in Claviceps paspali. II. Incorporation of labelled agroclavine, elymoclavine, lysergic acid and lysergic acid methyl ester. Acta Pharm. Suec., 3, 33-36. 

Naidoo, B., Cassady, J.M., Blair, G.E. and Floss, H.G. (1970) Biosynthesis of ergot alkaloids. Synthesis of chanoclavine-I-aldehyde and its incorporation into elymoclavine by Claviceps. J. Chem. Soc. Chem. Comm., 471—472. 

Ergot alkaloids, e.g. elymoclavine (7.55), are produced by a fungus, Clavicepspurpurea, which infects rye and other grasses. They have been extensively studied and there are several fascinating aspects of their biosynthesis. 

In considering the ring-closure of (7.53) to give (7.54) and the apparently connected double-bond isomerization, knowledge of the fate of the proton at C-9 in (7.53) is important. It was found to be incompletely retained ( 70%) during biosynthesis (that at C-10 is completely retained). In an experiment with a 1 1 mixture of [2- C]-and [4- H2]-mevalonate, the derived (7.53) showed the expected mixture of singly labelled species, but the elymoclavine (7.55) showed an appreciable percentage of double-labelled molecules, i.e. molecules which contained both C and deuterium. These results and the required double-bond isomerization are neatly accounted for in terms of the mechanism shown in Scheme 7.6 [30] in which there is an... 

C,9- H]Chanoclavine I was converted to elymoclavine with 70% tritium retention. (4R)-[2- " C,4- H]Mevalonate also gave elymoclavine and lysergic acid a-hydroxyethylamide with 70% tritium retention (Floss et ai, 1968). The percent tritium retention varied in the range 40-80% with the rate of alkaloid biosynthesis (Floss et al., 1974a). Low rates of alkaloid production correlated with low tritium retention at C(9). A mechanism was proposed to account for the loss of tritium (Fig. 6). The mechanism involves... 

Voigt and Zier (1970, 1971) proposed that the biosynthesis of ergot alkaloids could occur by the pathway dihydrochanoclavine I dihydro-agroclavine -> agroclavine elymoclavine. In experiments with ripening... 

With slant cultures of an Elymus-type Claviceps strain, Ohashi and Abe (1970) obtained cell-free conversion of agroclavine and elymoclavine to peptide alkaloids. There have been no reports so far of cell-free peptide alkaloid synthesis with shake cultures. Groger s laboratory (Maier et al., 1972) studied the activation reactions that could be involved in peptide alkaloid biosynthesis. They observed cell-free synthesis of lysergyl-CoA and activation of valine, serine, leucine, and proline. Activation was found in nonproducing, clavine alkaloid-producing, and peptide alkaloid-producing strains. The connection between these activities and peptide alkaloid synthesis is therefore uncertain. 

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