Mutants of Streptomyces cattleya producing N-acetyl and deshydroxy carbapenems related to thienamycin.

Rosi, D.; Drozd, Marion L.; Kuhrt, M. F.; Terminiello, L.; Came, Paul E.; Daum, Sol J.

doi:10.7164/antibiotics.34.341

Cited by 20 publications

(8 citation statements)

References 7 publications

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“…The variability of carbapenem side chain oxidation state as well as the discovery of a mutant strain6 of Streptomyces cattleya that produced deshydroxy thienamycin ( 10 ) instead of thienamycin ( 1 ) led us to believe that the thienamycin gene cluster7 encoded one or more enzyme(s) capable of oxidizing the C-2/C-6 moieties of carbapenems. Protein sequence analysis of ThnG and ThnQ indicated that each enzyme contained the Hx(D/E)x n H motif characteristic of non-heme Fe(II)/α-ketoglutarate (α-KG)–dependent dioxygenases, and these seemed promising candidates 8.…”

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Non-Heme Iron Oxygenases Generate Natural Structural Diversity in Carbapenem Antibiotics

et al. 2009

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Carbapenems are a clinically important antibiotic family. More than 50 naturally occurring carbapenam/ems are known and are distinguished primarily by their C-2/C-6 side chains where many are only differentiated by the oxidation states of these substituents. With a limited palette of variations the carbapenem family comprises a natural combinatorial library, and C-2/C-6 oxidation is associated with increased efficacy. We demonstrate that ThnG and ThnQ encoded by the thienamycin gene cluster in Streptomyces cattleya oxidize the C-2 and C-6 moieties of carbapenems, respectively. ThnQ stereospecifically hydroxylates PS-5 (5) giving N-acetyl thienamycin (2). ThnG catalyzes sequential desaturation and sulfoxidation of PS-5 (5), giving PS-7 (7) and its sulfoxide (9). The enzymes are relatively substrate selective but are proposed to give rise to the oxidative diversity of carbapenems produced by S. cattleya, and orthologues likely function similarly in allied streptomyces. Elucidating the roles of ThnG and ThnQ will focus further investigations of carbapenem antibiotic biosynthesis.

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Non-Heme Iron Oxygenases Generate Natural Structural Diversity in Carbapenem Antibiotics

et al. 2009

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“…However, ThnF is not likely to be the terminal step in thienamycin biosynthesis because an enzyme capable of deacetylating N-acetylthienamycin to thienamycin was found and purified from wild-type cultures of S. cattleya (33). The discovery of an S. cattleya mutant strain that solely produced N-acetylthienamycin further supports this hypothesis (34). Thus, a tentative biogenetic pathway to the cysteaminyl side chain from CoA can be proposed for thienamycin biosynthesis (Fig.…”

Section: Resultsmentioning

confidence: 92%

Four enzymes define the incorporation of coenzyme A in thienamycin biosynthesis

Freeman¹,

Moshos

Bodner

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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The enzymatic activities of three proteins encoded by the thienamycin gene cluster of Streptomyces cattleya (ThnR, ThnH, and ThnT) have been shown to incrementally cleave CoA to afford the active side-chain component of the ␤-lactam antibiotic thienamycin. These results supersede proposals based on earlier radiochemical incorporation experiments. For 20 years it has been thought that cysteine was directly incorporated into the antibiotic. Specific, stepwise truncation of CoA to 4-phosphopantetheine, pantetheine, and finally cysteamine was observed with ThnR, ThnH, and ThnT, respectively, in a series of coupled enzymatic assays. Pantetheinylated carbapenams were synthesized to address possible thienamycin biosynthetic intermediates and were shown to be effective substrates for the pantetheine-cleaving enzyme ThnT. Finally, a fourth gene, thnF, was shown to encode a protein capable of N-acetylating a model compound containing cysteamine in the presence of acetyl-CoA, consistent with the production of the S. cattleya cometabolite, N-acetylthienamycin. Taken together, these four enzymes are proposed to siphon CoA from primary metabolism to create the side chains for the predominant S. cattleya carbapenems, thienamycin and N-acetylthienamycin, in a process likely to be general for the broader class of these antibiotics.␤-lactam antibiotics ͉ carbapenem ͉ pyrophosphatase ͉ acylase ͉ phosphatase

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“…An intermediate in this process could also be methylated and successively modified to account for most of the chiral and side chain variations found in the carbapenems. The proposal of this biosynthetic scheme was greatly aided by studies with blocked mutants (Brown et al, 1977;Rosi et al, 1981). Nozaki et al (1984) also studied blocked mutants of S. griseus subsp.…”

Section: Carbapenemsmentioning

confidence: 99%