6-methylsalicylic acid biosynthesis

Fig. 70. Hypothetical scheme of 6-methylsalicylic acid biosynthesis E Enzyme SpH peripheric SH-group ScH central SH-group...

Formation of a polyketide involves condensation of acetyl-CoA with the appropriate number of malonyl-CoA units, modification of the completed poly-j3-ketone where required, and release of the product in stable form, as, for example, in 6-methylsalicylic acid biosynthesis (Scheme 3.4). The whole sequence occurs enzyme-bound, without release, or acceptance, of intermediates externally. Thus probing of the biosynthetic sequence by normal feeding experiments with possible intermediates fails. Instead, very properly, information is gained in different ways by working with the actual enzymes involved. The biosynthesis of several polyketides has been explored in this way, and most extensively that of 6-methylsalicyclic acid 3.14) [14-17]. 

S. antibioticus produces another unusual macrolide antibiotic [45] called chlorothricin (57) containing, in addition to the aglycone (modified methylsalicylic acid), saccharides, dideoxyhexoses in the first place. Their biosynthesis was investigated by the incorporation of stable isotopes [45-47]. The compounds were only active in a synthetic medium and inactive in a complex one [48]. Compounds designated MC031-034 (58-61) are similar to chlorothricin mentioned above and were isolated from the cultivation broth of Streptomyces sp. collected in Japan. Another compound, 2-hydroxychlorothricin (62), with antitumor activity, was isolated [66] from Streptomyces K818. 

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]. 

In 1955, 6-methylsalicylic acid was the first fungal metabolite to be used to test the acetate hypothesis for the biosynthesis of aromatic compounds. The original proposal had been made by Collie in 1907 and then developed by Birch in 1953. This experiment, like many early biosynthetic studies, was carried out by... 

On the Role of 6-Methylsalicylic Acid in the Biosynthesis of Fungal Benzoqui-nones. Acta Chem. Scand. 20, 151 (1966). 

Aromatic biosynthesis, aromatizatioa biosynthesis of compounds containing the benzene ring system. The most important mechanisms are 1. the shi-kimate/chorismate pathway, in which the aromatic amino acids, L-phenylalanine, L-tyrosine and L-trypto-phan, 4-hydroxybenzoic acid (precursor of ubiquinone), 4-aminobenzoie acid (precursor of folic acid) and the phenylpropanes, including components of lignin, cinnamic acid derivatives and flavonoids are synthesized and 2. the polyketide pathway (see Polyke-tides) in which acetate molecules are condensed and aromatic compounds (e.g. 6-methylsalicylic acid) are synthesized via poly-fl-keto acids. Biosynthesis of flavonoids (e.g. anthocyanidins) can occur by either pathway. 

Examples of P. are Tetra clines (see), Griseofulvin (see), Macrolide antibotics (see), Cydoheximide (see), and various fungal products such as orsellinic acid, 6-methylsalicylic acid and cyclopaldic add. [ The Biosynthesis of Acetate-Derived Phenols (Polyketides) by N.M.Packter pp. 535-570, in The Biochemistry of Plants, V6I4, 1980 (Edit. P.K. Stumpf), Academic Press S.Sahpaz et al. Phytochemistry 42 (1996) 103-107]... 

Some of the bound keto groups may be reduced by pyridine nucleotide-dependent dehydrogenases (C 2.1, see the formation of 6-methylsalicylic acid, D 3.3.1), and some of the activated CHg-groups may be alkylated by S-adenosyl-L-meth-ionine (C 3.3, see the biosynthesis of tetracyclines, D 3.3.7), or may be substituted by dimethylaUyl pyrophosphate (D 6), see the formulas of lupulone and humulone, the bitter principles of hop cones used in brewing beer (F 1). 

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 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 bacterial biosynthesis of 6-methylsalicylic acid (6-MSA) (4) has been studied in detail and the administration of radioactive malonate to Mycobacterium phlei has provided 6-MSA with a labelling pattern consistent with its derivation by the acetate-polymalonate biosynthetic route. Some activity from malonate was incorporated into the acetate-derived C-methyl group and this partial conversion of malonate into acetate has been recorded previously in fungal studies. Related studies on the biosynthesis of 6-MSA and salicylic acid by Mycobacterium fortuitum have shown that acetate was much more efficiently incorporated into the former compound. It has been concluded, therefore, that these structurally similar phenolic acids are derived by different biosynthetic pathways. 

Salicylic Acid.—It has recently been demonstrated (see last year s Report ) that whereas 6-methylsalicylic acid in both micro-organisms and higher plants is acetate-derived, salicylic add itself is formed in both groups of organisms by the shikimic add pathway. Further confirmation of this has come from Marshall and Ratledge, who have studied the enzymology of biosynthesis of salicylic acid in Mycobacterium smegmatis. These authors isolated the enzyme salicylate synthetase, which catalyses the last step (see Scheme 1) in synthesis of salicylic acid (1) from isochorismic add (2), formed in turn from chorismic (3) and shikimic adds (4). The enzyme has no cofactor requirements and converts (2) directly into (1). No evidence could be obtained for the presence in bacterial cultures of the possible intermediate 2,3-dihydroxy-2,3-dihydrobenzoic add. 

L Abell, C., MJ. Garson, FJ. Leeper, and J. Stauton Biosynthesis of the Fungal Metabolites Altemariol, Mellein, Rubrofusarin, and 6-Methylsalicylic Acid from Acetic Acid-2,2,2-d3. Chem. Commun. 1982,1011. 

Monodictyphenone has been previously isolated from a marine fungus Monodictys putredinis as well as an engineered strain of A mdidans. This strain of A. nidulans expressed the (3area lozoyensis polyketide synthase gene involved for 6-methylsalicylic acid (15) biosynthesis, and the authors could not determine whether the monodictyphenone produced in addition to 6-methydsalicylic acid was due... 

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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