Cyclopenin

Early biosynthetic studies involving feeding experiments with [cflrfcoxy- C]anthr anilic acid, DL-[l- C]phenylalanine, DL-[2- C]phenylalanine, DL-[3- C]phenylala 

THREE-MEMBERED HETEROCYCLIC RINGS AND THEIR FUSED DERIVATIVES 

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Benzodiazepine Bases.—It is known that phenylalanine is an intact precursor for the benzodiazepine bases cyclopenin (124) and cyclopenol (125), via (122) and (123), in Penicillium cyclopium cultures.102 Examination of the stereochemical course of the necessary tritium loss from C-3 of the precursor amino-acid on formation of (124) has given a surprising result more than half of the tritium was... 

Significant progress has been made in the identification of enzymes from P. cyclopium which catalyse the various reactions of cyclopenin (124) and cyclopenol (125) biosynthesis.105 One of these is cyclopeptine dehydrogenase which catalyses the interconversion of (122) and (123).105a,1°6 Further research107 has shown that... 

The first correct structures of benzodiazepine alkaloids, namely, those of cyclopenin (3) and cyclopenol (4), were published in 1963 by Mohammed and Luckner (47). In 1965 the structure of the antibiotic anthramycin (8) was established (28), followed by tomaymycin (12) and oxotomaymycin (13) in 1971 (26). The biosynthetic pathway to both the cyclopenin group and the anthramycin-tomaymycin group of alkaloids was elucidated in the following years and has been included in previous reviews (32,33). Later additions to the small list of naturally occurring benzodiazepines comprise aszonalenin (6) (16), asperlicin (7) (31), and auranthine (5) (85). The biosynthesis of the latter alkaloids has not yet been investigated in detail. 

Table II compiles melting points, optical rotations, and methods used for the structural elucidation of the known benzodiazepine alkaloids. Intensive studies of the degradation of the molecules as well as attempts at total synthesis have been undertaken only with members of the cyclopenin and the anthramycin-tomaymycin groups. In the following a short summary is given of the reactions and intermediates that provided essential information about the structures of the basic skeletons of the alkaloids.

Cyclopenin and precursor alkaloids Penicillium cyclopium Westling" 5,7,18,47... 

Fig. 2. Degradation of cyclopenin and cyclopenol (45,51). Degradation products are numbered in relation to the parent alkaloids. 

These data, taken together with mass spectrometric degradation and other spectral evidence (see Table II), clearly favor a seven-membered cyclic peptide formed from anthranilate and phenylalanine as the basic skeleton of cyclopenin (3) and cyclopenol (4). This structure, including the epoxide linkage, was further supported by biosynthetic probing (51,52) (see Section IV,A). 

The first total synthesis of cyclopenin (3) by Rapoport and colleagues (45,71) (Fig. 3) started with 2-amino-A -methylbenzamide, which was reacted with... 

All naturally occurring benzodiazepines are biosynthesized from anthranilic acid. The benzodiazepine moiety is formed by reacting this central precursor with derivatives of phenylalanine (cyclopenin group), tyrosine (tomaymycin-anthramycin group), glutamine (auranthine), or tryptophan plus another molecule of anthranilic acid (asperlicin). Intensive biosynthetic work has been published for the cyclopenin and tomaymycin-anthramycin groups only. 

These findings prove the pathway of cyclopenin-cyclopenol biosynthesis shown in Fig. 5. In the first step, the cyclic dipeptide cyclopeptine (1) is formed... 

Fig. 5. Biosynthetic pathway to cyclopenin and cyclopenol (37). Enzyme activities involved (I) cyclopeptine synthetase (2) cyclopeptine dehydrogenase (3) dehydrocyclopeptine epoxidase (4)...

The biosynthetic scheme (Fig. 5) suggests the involvement of four enzymes in the formation of cyclopenin. All of these activities have been characterized in more detail (for an early summary, see Ref. 34). 

A mechanism for the transformation of the benzodiazepine to the quinoline structure, which accounts for all data obtained from both cyclopenase- and acid- or base-catalyzed reactions, has been proposed by White and Dimsdale (79) (Fig. 8). The conversion is induced by attack of an electron acceptor (e.g., H" ) at the epoxide oxygen or by attack of an electron donor (e.g., OH ) on the N-1 atom. Bond formation between C-10 and C-5a of cyclopenin is facilitated by the close approach of these two atoms in the boat conformation. Therefore, the formation of a tricyclic intermediate appears likely, although a compound of this type has not been isolated so far. Another mechanism that would explain the enzymatic and acid-catalyzed conversions was proposed by Luckner and Never (42). 

Fig. 8. Possible mechanism of the conversion of cyclopenin (3) to viridicalin (15), triggered by an electrophilic attack at the epoxide group (S3, according to Ref. 79).

Fig. 10. In vitro activities of the enzymes of alkaloid biosynthesis and rates of alkaloid formation in hyphae of Penicitlium cyclopium (75). Cultivation conditions were as described in the legend to Fig. 9. At the times indicated by arrows, cycloheximide (100 p.g/ml) was added to the culture medium. All values are in units per 1 cm of culture area. (A) Anthranilate adenylyltransferase (AA) (100 = 5.6 pkat) ( ) cyclopeptine dehydrogenase (CD) (100 = 40 pkat) ( ) dehydrocyclopeptine epoxidase (DE) (100 = 0.42 pkat) (A) cyclopenin m-hydroxylase (CH) (100 = 12 pkat) (O) cyclopenin-cyclopenol formation in vivo (100 = 9 pmol/sec).

A characteristic and exclusive constituent of the conidiospores is the enzyme cyclopenase, which catalyzes the conversion of the benzodiazepines cyclopenin-cyclopenol to the quinolines viridicatin-viridicatol (see above). This enzyme... 

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