Aspergillus amstelodami

C NMR data for a substance isolated from Aspergillus amstelodami are given in [343]. (216) H NMR parameters for eight metabolites from... 

Extracts from Aspergillus amstelodami catalysed the formation of cyc/o-L-prolyl-2-(l,l-dimethylallyl)-L-tryptophanyl (122) from [1-3H]DMAPP and cyc/o-L-prolyl-L-tryptophanyl (123).389 The same enzyme could isoprenylate cyclo-L-alanyl-L-tryptophanyl, but cyc/o-pentylidene-ethyl pyrophosphate did not function as a substrate. The conclusion that echinulin (124) is derived from the prenylation of a pre-formed cyc/o-analyltryptophanyl system is further supported by the isolation of (125) from the mycelium of A. amstelodami,390 whilst the co-occurrence and biogenetic relationship between (125), (12b), (127), and (128) has led to speculation concerning the route to neoechinulin. 

Miscellaneous Indole Alkaloids.—Echinulin (38) is known to be assembled from tryptophan, mevalonic acid, and alanine.24-26 The indole derivative (37) has now been shown27 to be converted efficiently by Aspergillus amstelodami into echinulin and is, therefore, probably an intermediate. 

Echinulin.— Most convincing evidence has been obtained that (126) is an intermediate in the biosynthesis of echinulin (127) in Aspergillus amstelodami Possible mechanisms for introduction of the isoprene units on to the benzene ring of echinulin (127) and neoechinulin (128) are circumscribed by the results of feeding... 

Mould Metabolites.—In the search for later intermediates in the biosynthesis of echinulin and neochinulin three new prenylated indole derivatives have been isolated from the mycelium of Aspergillus amstelodami these are neoechinulin A (19), neoechinulin B (20), and neoechinulin C (cryptoechinulin A) (21). The isolation of neoechinulin A lends credence to the proposal that in the biosynthesis of echinulin the first isopentenyl group is introduced to position 2 of a preformed cyc/o-alanyltryptophan system, while the isolation of (19)—(21) suggests that neoechinulin (22) may be formed by a dehydrogenation followed by oxidative fission of the alanyl methyl group (CHMe —> C=CH2 C=0). ... 

Brevianamide.—Echinulin (109) is elaborated in Aspergillus amstelodami along a pathway which includes (110) and (111). The structurally related metabolite, brevianamide A (112), isolated from Penicillium brevicompactum, appears to be derived in a similar way. Radioactive mevalonate, proline, and tryptophan gave labelled brevianamide A. Incorporation of cyclo-L-[methylene- C]-tryptophyl-L-[5- H]proline [as (113)] without change in isotope ratio indicated that this precursor was utilized intact for brevianamide A production. As further evidence of its role in brevianamide A biosynthesis (113) has been isolated from P. brevicompactum cultures. By analogy with echinulin biosynthesis, (114) could lie between (113) and brevianamide A (112). Its isolation" (from A. ustus) provides support for this suggestion. 

Labeled samples of leucine 205 were fed to Aspergillus amstelodami to produce the metabolite echinulin 210. The A-pro-S methyl group of leucine was shown to be incorporated into the (Z)-methyl groups of the isoprenoid residues via catabolism to HMG-CoA (hydroxymethylglutaryl-coenzyme A) (see Section VIII.E) and mevalonic acid (181). 

Mould Metabolites.— The extensive work of Kishi and his collaborators on the metabolites of Aspergillus amstelodami has been published in detail, regrettably in a Journal not readily intelligible to most Western readers. This work includes the total synthesis of echinulin, " previously reported in brief,and a total synthesis of neoechinulin (29) (Scheme 5). The crucial indole derivative... 

Echinulin is a triisoprenylated cyclic dipeptide isolated from Aspergillus amstelodami and Aspergillus echinulatus (Fig. 12) [52]. Birch et al. established that the biosynthetic precursors to echinulin are L-tryptophan, L-alanine, and mevalonic acid [53] by feeding radioactive precursors to Aspergillus amsteloda-mu In these experiments it was found that [1- C] sodium acetate, dl-[1- C]-alanine,DL-[l- C]-tryptophan, [2- C]-mevalonic acid lactone, and [1- C]-glycine were incorporated into echinulin with incorporation rates of 4.25 %, 0.12 %, 1.36%, 0.74%, and 0.84%, respectively (Scheme 31). 

In addition, it was further established by Slater et al. that cyc/o-L-alanyl-L-tryptohan are precursors to echinulin in vivo (Scheme 32) [54]. In this instance, the radioactive precursor cyc/o-L-alanyl-L-[met/2y/ene- C]-tryptophan was fed to cultures of Aspergillus amstelodami and gave echinulin with high levels of incorporation (9-16%). 

A similar experiment was later reported by Marchelli et al. [55], wherein doubly labeled cydo-L-[U- C]alanyl-L-[5,7- H2]tryptophan and cyc/o-L-[U- C] alanyl-D-[5,7- H2] tryptophan were synthesized and fed to cultures of Aspergillus amstelodami. A series of echinulins were isolated and showed incorporations as follows neoechinulin A (88%), neoechinulin B (43%), neoechinulin C (43%), neoechinulin D (34%), and neoechinulin (43%) respectively, relative to the radioactivity recovered in echinulin. The L,D-cyclic dipeptide was very inefficiently incorporated into the natural metabolites thus further confirming cyclo-L-alanyl-L-tryptophyl as a key biosynthetic precursor to this family of natural substances. 

In 1973, Allen demonstrated that the monoprenylated cyclic dipeptide cyclo-L-alanyl-2-(l,l,-dimethylallyl)-L-tryptophan was incorporated into echinulin in cultures of Aspergillus amstelodami [56]. In this study, the reverse prenyiated cyclic dipeptide was doubly labeled with and by treating a partially purified prenyl transferase obtained from Aspergillus amstelodami with cyclo-L-Ala-L-[3- C]-Trp and [l- H]-3-methyl-2-butenyl 1-pyrophosphate (Scheme 33)... 

As shown in Scheme 34, [4,6- H2]-tryptophan and [5,7- H2]-tryptophan were synthesized and fed to cultures of Aspergillus amstelodami. The [5,7- H2]-tryptophan was incorporated into echinulin and neoechinulin B with 2% and 103% retention of tritium activity, respectively. The [4,6- H2l-tryptophan was incorporated into echinulin with 102% retention of tritium activity and into neoechinulin with 48% loss of tritium activity. These experiments are complementary and clearly demonstrate that the introduction of the isoprene units go via a direct electrophilic aromatic substitution reaction mechanism. It shoidd also be noted that Fuganti et al. isolated cryptoechinulin from Aspergillus amstelodami during the course of their biosynthetic work on echinulin [58]. 

The stereochemistry of the a,j0-desaturation of cryptoechinulin was reported by Fuganti et al. and is summarized in Scheme 35 [59]. These workers prepared L-tryptophan stereospecifically labeled with tritium in the -methylene position from labeled serine, indole, and fibre-entrapped tryptophan synthetase obtained from E. coli. When (3 i )[3 - H 3 - C]-L-tryptophan was fed to Aspergillus amstelodami, incorporation into echinulin and cryptoechinulin took place with 95% and 98% retention of tritium activity, respectively. Feeding of (3 S)[3 - H 3 - C]-L-tryptophan to Aspergillus amstelodami gave incorporation into echinulin and cryptoechinulin with 96% and 5% retention of tritium activity, respectively. Thus, in the desaturation reaction the pro-S hydrogen is stereospecifically removed. 

Barrow et al. investigated the stereochemistry of the C-C bond-forming reaction in the aromatic isoprenylation in echinulin biosynthesis utilizing [(5i )- Hj and [(5S)- H]-mevalonates [60]. In addition, feeding of [1,2- 2]-acetate to Aspergillus amstelodami showed that the ( )-methyl groups in the isoprene moieties are derived only from C-2 of mevalonic acid (Scheme 36). The ( )-methyl group was found to be enriched but not coupled to the adjacent olefinic center. 

Grundon et al. examined the possibility that the reverse prenyl unit was introduced via an indolic N-prenylated precursor followed by aza-Claisen rearrangement and 1,2-migration as shown in Scheme 38 [61]. These workers synthesized l-([l- H]-3,3-dimethylallyl)-L-tryptophan and cyclo-L-alanyl-l-([l- H]-3,3-dimethylaUyl)-L-tryptophan and fed these labeled substances to cultures of Aspergillus amstelodami and found that neither compound was incorporated in radiochemically significant amounts into echinulin. 

In order to confirm the above pathway, Fuganti et al. performed a second experiment using a doubly labeled leucine derivative to examine the fate of positions 2 and 3 of leucine (Scheme 42) [64]. In the first experiment, (4S)-[5- ]-l-leucine admixed with (4S)-[5- C]-L-leucine was fed to cultures of Aspergillus amstelodami yielding echinulin with 30% loss of tritium. In a second experiment, (4S)-[5- H]-L-leucine admixed with [2- C]-L-leucine ratio =... 

Cardillo, R., C. Fuganti, D. Ghiringhelli, and P. Grasselli Stereochemical Course of the a,P-Desaturation of L-Tryptophan in the Biosynthesis of Crypto-echinuline A in Aspergillus Amstelodami. J. Chem. Soc. Chem. Comm. 1975, 778. 

Marchelli, R., A. Dossena, A. Pochini, and E. Dradi The Structures of Five New Didehydropetides related to Neochinulin, Isolated from Aspergillus Amstelodami. J. Chem. Soc., Perkin Transactions I 1977, 713. 

Aspergillus amstelodami Eleagnus angustifolia Claviaeps sp. (strain SD-58) Clavioeps purpurea Claviaeps purpurea Papaver sormiferum... 

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