Cyclopeptine

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... 

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)...

Fio. 6. Activities of the cyclopeptine synthetase complex (19). (1) Anthranilic acid ade-nylyltransferase (2) L-phenylalanine adenylyltransferase (3) covalent binding of anthranilic acid (4) covalent binding of L-phenylalanine (5) methylation of enzyme-bound L-phenylalanine by 5-ade-nosyl-L-methionine (6) translocation of AI-methyl-L-phenylalanine to enzyme bound anthranilic acid (7) release of cyclopeptine by cyclization. 

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).

Feeding experiments with various possible intermediates have shown that the two N-methyl cyclic peptides cyclopeptine (160) and its dehydroderivative (161) are precursors for cyclopenin and cyclopenol. Further, (160) and (161) could be isolated from the culture and were themselves labelled by radioactive phenylalanine and anthranilic acid. In addition, cyclopeptine (160) was found to be reversibly transformed into dehydrocyclopeptine (161). Negative results with compounds likely to be implicated earlier on the pathway suggested that the formation of cyclopeptine (160) from the precursor amino-acids took place with enzyme-bound intermediates. The pathway established so far is summarized in Scheme 16. 

The enzymes involved in the alkaloid metabolism of P. ovoloplum become measurable during the transition from tne trophophase to the idiophase. The activity increase of cyclopeptine dehydrogenase, dehydrocyclopeptine ep-oxidase and cyclopenin m-hydioxylase is inhibited by 5-fluorouracil and cycloheximide, indicating that it depends on RNA and protein biosynthesis (7 - 9). Hence like many other secondary metabolic activities (cf, 2,... 

Winter, S. Werner and M. luckner, Cyclopeptin und Dehydrocyclopeptin, Zwischenprodukte der Biosynthese von Alkaloiden der C clopenin-Viridicatin-Gruppe bei Penicillium cyclopium WESTIIRG, Europ. J. Biochem. 37, 78 - 85 (l973). 

Figure 3.61 Biosynthetic pathway to cyclopenin and cyclopenol, and their conversion to viridicatin and viridicatol, respectively, in Penicillium cyclopium. (i) cyclopeptinsynthase complex (ii) cyclopeptin dehydrogenase (iii) dehydrocyclopeptin epoxidase (iv) cyclopenin m-hydroxylase (v) cyclopenase.

Enzymes of alkaloid biosynthesis are induced in a coordinated manner by an internal signal during development of the mold. However, the rates of alkaloid production increase later than enzyme activities, The increase of both processes can be stopped by addition of inhibitors of gene expression, e.g., 6-fluorouracil or cycloheximide (cf. arrows), indicating that alkaloid production is controlled by the formation of a protein which is not an enzyme of alkaloid biosynthesis. Ordinate relative values A A anthranilic acid adenylyl transferase (a) CD cyclopeptine dehydrogenase (v) DE dehydrocyclopeptine epoxidase ( ) CmH cyclopenin m-hydroxvlase (A)... 

Benzodiazepine Alkaloids and 4 Phenylquinolines (Figs. 154 and 155). The benzodiazepine alkaloids formed in moulds of the genus Penicillium are cyclic peptides formed from anthranilic acid and L-phenylalanine. The key intermediate cyclopeptine is probably synthesized by a polyfunctional protein resembling... 

Cyclopeptine synthetase 2 cyclopeptine dehydrogenase 3 dehydrocyclopeptine epoxidase 4 cyclopenin m-hydroxylase 5 cyclopenase... 

The benzodiazepines, e.g. cyclopenin (7.46), are, formally at least, related to the diketopiperazines in being formed from two amino acids. For cyclopenin (7.46) and cyclopenol (7.47), in Penicillium eye-lopium, they are anthranilic acid (7.43) and phenylalanine (7.17). The first detectable intermediates are cyclopeptine (7.44) and dehydrocyc-lopeptine (7.45), which are interconvertible earlier intermediates apparently only exist enzyme-bound, not being equilibrated with fed precursors. 

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