Selection of a specifically blocked mutant of. Streptomyces cinnamonensis: Isolation and synthesis of 26-deoxymonensin A.

Abstract

Streptomyces cinnamonensis produces the polyether ionophore antibiotic monensin A. Following a single round of mutagenesis by UVlight, a derivative of this strain has been isolated, which secretes a new metabolite identified as 26-deoxymonensin A (3). The structural elucidation of the newmetabolite followed from a spectroscopic analysis, and its identity was proven conclusively following a comparison to 26-deoxymonensin A (3) obtained synthetically from monensin A. The preparation of labelled forms of 3 is described, together with incorporation experiments using the parent strain of S. cinnamonensis. Only very low levels of incorporation of 3 into monensin A were observed.Monensin A (1) is an important polyether ionophore antibiotic, produced13 by Streptomyces cinnamonensis. The biosynthesis of this antibiotic has attracted a great deal of interest2~5), following proposals3»4) that the cyclic ether groups in monensin A might arise by the novel mechanism shown in Scheme1, proceeding via a cascade of cyclisation steps on a triepoxide intermediate. This route is supported by the results of isotope labelling experiments3~5) that define the biosynthetic origins of the carbon and oxygen atoms in monensin A, and similar experiments with other polyethers, including narasin6), lenoremycin7), lasalocid A8), ICI 1396039) and maduramicin10), strengthen a belief that this very appealing concept may be applied generally10 to account for the formation of cyclic ether rings in manyof the other knownmembersof this family of secondary metabolites.In order to further characterize the pathway to the monensins A (1) and B (2) in S. cinnamonensis, we have generated mutants blocked in monensin production, and describe here the isolation of one such mutant that accumulates mainly 26-deoxymonensin A (3); the mutant must carry a lesion in the hydroxylation step occuring at the terminal C-26 position. The structure assigned to 3 has been confirmed by direct comparison with a sample of 26-deoxymonensin A prepared chemically from monensin A.There have been no other reports, to date, on the isolation of mutants specifically blocked in the monensin biosynthetic pathway. Also, no hona fide biosynthetic intermediates have been isolated, although 3-0-demethylmonensins A (4) and B have been recovered12-* as minor components from the fermentation broth of a normal producing strain.

Intermediates in monensin biosynthesis: A late step in biosynthesis of the polyether ionophore monensin is crucial for the integrity of cation binding

“…The spectroscopic properties of the purified 3- O -demethylmonensin A were in complete agreement with those previously reported for this compound, isolated as a minor component of a wild-type S. cinnamonensis strain [23]. Similarly, the spectroscopic properties of the isolated 26-dehydroxymonensin A were identical with those described for this compound obtained either from a specifically-blocked mutant of wild-type S. cinnamonensis [24] or from a wild-type fermentation conducted in the presence of the potent methyltransferase inhibitor metapyrone [25]. …”

Production of 26-deoxymonensins A and B by Streptomyces cinnamonensis in the presence of Metyrapone

“…By using metyrapone and methylation inhibitors (aminopterin and sulfonamides), we obtained corresponding intermediates in reasonable yields. Nevertheless, the roles of 3-0-demethylmonensins and 26-deoxymonensins in normal monensin biosynthesis remain unsettled (2,10).…”

Cloning, sequencing, and expression of the gene encoding methylmalonyl-coenzyme A mutase from Streptomyces cinnamonensis

“…S. cinnamonensis A3823.5 (Eli Lilly) was used as the source strain and was cultured as previously described (2). S. lividans TK64, employed for the propagation of Streptomyces plasmids and as the expression host, was cultured and transformed according to the method of Hopwood et al (15).…”