Patulin biosynthesis

Paterson, R. R. M. (2004). The isoepoxydon dehydrogenase gene of patulin biosynthesis in cultures and secondary metabolites as candidate PCR inhibitors. Mycol. Res. 108, 1431-1437. 

Murphy, G., G. Vogel, G. Krippahl, and F. Lynen Patulin Biosynthesis. Role of Mixed-Function Oxidases in the Hydroxylation of m-cresol. Eur. J. Phytochem. 49, 443 (1974). 

Scott and co-workers (1967, 1973) have used deuterated precursors to delineate patulin biosynthesis. The method involves preparing deuterated putative intermediates, feeding them to the fungi, and isolating the enriched products. The compounds are then purified by the usual organic methods and analyzed by mass spectrometry. The advantages of this technique are (a) it is much easier to synthesize precursors with rather than (b) mass... 

Scheme 1. Proposed pathway of patulin biosynthesis. This figure is modified after the various experimental results obtained from different research groups (Zamir, 1973 Scott et ai, 1973 Light, 1969 Light and Vogel, 1975 Murphy et al., 1974 Scott and Beadling, 1974 Murphy and Lynen, 1975 Forrester and Gaucher, 1972).

Most of the steps in Scheme 1 have been established at the enzymatic level (heavy arrows the enzymes detected are shown above these arrows). The unanswered questions in the biosynthesis of patulin are the following (a) What is the absolute configuration of the optically active deuteropatulin generated from the feeding of ring-deuterated aromatic precursors (b) Is deoxypatulinic acid (37) in Scheme 1 involved in patulin biosynthesis or in its metabolism (c) Is patulin-lactone (35) a metabolite of P. patuluml If not, why are the side-chain deuteriums lost in the conversion of the aromatic precursors into patulin ... 

Fig. 36. Intermediacy of phyllostine in patulin biosynthesis, modified after Sekiguchi and Gaucher (1978).

Cell-free systems capable of in vitro synthesis of 6-methylsalicylic acid (6-MS A) and a related tetraketide, orsellinic acid, were developed long before the advent of recombinant DNA technologies in the field of natural product biosynthesis [113-115] (Fig. 5). Since then, the biosynthetic mechanisms and molecular recognition features of 6-methylsalicylic acid synthase (6-MSAS) have been extensively studied. 6-MSAS initiates synthesis with an acetyl group derived from acetyl Co A, extends the polyketide chain to a tetraketide via three decar-boxylative condensations of malonyl CoA-derived extender units, and uses NADPH to specifically reduce one of resulting carbonyls to a hydroxyl group. In its natural producer, Penicillium patulum, the product, 6-MSA is subsequently glycosylated to form the antibiotic patulin [116]. 

A step in the biosynthesis of PATULIN (17) has been clarified by the finding that meta hydroxy benzyl alcohol is hydroxylated to yield gentisyl alcohol which in turn is converted to patulin via the aldehyde 55. 

Two interesting metabolites of Penicillium patulum are patulin and penicillic acid. Their biosynthesis involves the cleavage of an aromatic ring. These substances are mycotoxins and their activity in this context is discussed in Chapter 9. 

The biosynthesis of patulin was also studied using 02]acetate and 2 gas and then measuring the oxygen-18 induced chemical shifts in the NMR spectrum to locate the site of the This showed that only the carbonyl oxygen was derived from acetate and that the others were derived by oxidative processes. Many of the enzyme systems that mediate these steps have been isolated from P. patulum. 

A compound that serves as an intermediate in the biosynthesis of patulin (6), phyllostine (also known as epoxydon) (55), (Fig. 5.7) has been identified as a phytotoxin from a Phyllosticta species that is pathogenic to red clover (Ballio, 1981). Patulin also has been reported to possess phytotoxic activity (Stoessl, 1981). Further, several structurally related compounds from a number of different fungal species seem to be involved in phytotoxicity (Stoessl, 1981). 

Zamir, L. O., The biosynthesis of patulin and penicillic acid, in The Biosynthesis of Mycotoxins (P. Steyn, ed.), 223-268, Academic Press, New York, 1980. 

The biosynthesis of the aromatic metabolite, 6-methylsalicylic acid [3.14), has been discussed above. This acid is the source of a variety of metabolites in which hydroxylation and oxidation of the aromatic nucleus and side-chain methyl group occur in major pathways [4] following decarboxylation to m-cresol [3.42). A terminus in this set of oxidative reactions is patulin [3.45). 

The relatively high incorporations of tracer molecules into secondary metabolites in micro-organisms has enabled detailed studies to be made of the biosynthesis of the fungal metabolites patulin (10) and multicolic acid (79). Carbon-14 work has demonstrated the polyketide origin of patulin, and the biosynthesis of multicolic acid from acetate has been studied by C-n.m.r. spectroscopy. The biosynthesis of patulin has been investigated extensively at the enzymology level. In the first... 

The fungal metabolites patulin (10) and multicolic acid (79) are derived in nature via oxidative cleavages of polyketide derived aromatic intermediates a similar pathway has been established for the tetronic acid penicillic acid (185). By contrast, the biosynthesis of carolic acid (76a) which is found with dehydrocarolic acid (75) in P. charlesii, has been shown to occur from Krebs cycle intermediates. 

The biosynthesis of patulin (10) has been studied extensively, since the molecule represents a relatively simple model system in which to examine the detailed enzymology of polyketide biosynthesis. Patulin is biosynthesized by the fungus Penicillium patulum via an oxidative pathway from 6-methylsalicylic acid (188) which is synthesized from acetyl-CoA and malonyl-CoA. The major pathway from (188) and the biosynthetic relationships of the phenolic secondary metabolites of P. patulum are... 

The biosynthesis of the fungal tetronic acids penicillic acid (185) and carolic acid (76 a), which are closely related structurally to patulin and multicolic acid, have also been studied extensively. Early work with " C-labelled precursors has clearly demonstrated the polyketide origin of penicillic acid, and also the intermediacy of orsellinic acid... 

The biosynthesis of the fungal tetronic acid carolic acid (76 a) in Penicillium charlesii is markedly different from that of patulin, multicolic acid and penicillic acid. Carbon-14 work has demonstrated that the carbon sub-unit C-3, C-4, C-9 in the acid is derived from a C-4-dicarboxylic acid such as succinate, whereas the remaining six carbons (C-1, C-2, C-5, C-6, C-7, C-8) are derived from two malonate units (C-5, C-6, C-7, C-8) and just one acetate unit (C-1, C-2) (Scheme 19) (775). It is proposed then that the biosynthesis of carolic acid proceeds from a C4-dicarboxylic acid first to either y-carboxymethyltetronic acid (196) or to carlosic acid (77 b) hydroxylation of carlosic acid then gives rise to carlic acid (77 a) which on decarboxylation produces carolic acid 176). 

Scott, A. L, and L. Beadling Biosynthesis of Patulin. Dehydrogenase and Dioxygenase Enzymes of Penicillium patulum. Bioorganic Chem. 3, 281 (1974). 

Patulin. Scott and his collaborators have published full details of their work on the biosynthesis of patulin (157), an antibiotic isolated from cultures... 

Evidence that acetyl-CoA is the actual active acetate in fungal aromatic biosynthesis was provided by Bassett and Tanenbaum (i960), who showed (a) that coenzyme A could be isolated from the mycelium of P. patulum (b) that radioactive acetyl-CoA, synthesized either from fungal or yeast coenzyme preparations, could be transformed into radioactive patulin by a cell-free extract obtained by the dilute ammonia treatment of mycelial mats. It was also found that biosynthetically-labelled 6-methylsalicylate could be enzymically rearranged to patulin by such extracts. Use of the ammonia extractive procedure followed by partial ammonium sulfate fractionation, gave Lynen and Tada (I96I) an enzyme mixture which synthesized radioactive 6-methylsalicylate from acetyl-1- C-CoA,... 

Tanenbaum, S. W., and E. W. Bassett The biosynthesis of patulin. I. Related aromatic substances from Penicillium patulum, strain 2159 A II, the general physiology of several strains of P. patulum. Biochim. et Biophys. Acta 28, 21, 247 (1958b). 

The secondary biosynthesis of the mycotoxin patulin, as described in detail in Chapter 7, begins with the formation of 6-methylsalicylic acid, and the regulation of this first step is therefore rate-limiting overall. However, it is the control of further steps beyond 6-methylsalicylate that determines the extent to which patulin, or other products of the pathway, actually accumulate (Bu Lock et al., 1965). 

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