Summary auto-generated
This study investigated the role of orf2* (now designated tylM2) in tylosin biosynthesis by Streptomyces fradiae, a soil bacterium that produces the macrolide antibiotic tylosin. Researchers used targeted gene disruption and bioconversion experiments to characterize this gene. The orf2* product is a glycosyltransferase that adds mycaminose, the first sugar in tylosin's structure, to the polyketide precursor tylactone. Surprisingly, disruption of orf2* not only eliminated glycosylated tylosin products but also prevented tylactone accumulation under normal fermentation conditions. When the gene was reintroduced, tylosin production recovered at only 10% of wild-type levels. However, when researchers fed glycosylated tylosin precursors to the disrupted strain, tylactone accumulated, indicating that glycosylated macrolides stimulate polyketide metabolism. The results suggest that orf2* disruption may create a polar effect on downstream genes, particularly orf4* (ccr), which encodes crotonyl-CoA reductase that supplies building blocks for polyketide synthesis. This work reveals an unexpected regulatory connection between glycosylation and polyketide metabolism in tylosin production.
Key findings
- Orf2* encodes mycaminosyltransferase (TylM2), which adds the first sugar (mycaminose) to the tylosin polyketide aglycone
- Disruption of orf2* prevented both glycosylated tylosin production and tylactone accumulation, suggesting polyketide metabolism is impaired without glycosylation
- Glycosylated tylosin precursors stimulated polyketide metabolism and tylactone accumulation in the orf2*-disrupted strain, indicating positive feedback from glycosylated macrolides
- Re-introduction of orf2* restored tylosin production to only ~10% of wild-type levels, possibly due to polar effects on downstream genes like orf4* (ccr)
- The study demonstrates an unexpected regulatory connection between glycosylation and polyketide biosynthesis in Streptomyces fradiae
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Abstract
Three glycosyltransferases are involved in tylosin biosynthesis in Streptomyces fradiae. The first sugar to be added to the polyketide aglycone (tylactone) is mycaminose and the gene encoding mycaminosyltransferase is orf2* (tylM2). However, targeted disruption of orf2* did not lead to the accumulation of tylactone under conditions that normally favour tylosin production; instead, the synthesis of tylactone was virtually abolished. This may, in part, have resulted from a polar effect on the expression of genes downstream of orf2*. particularly orf4* (ccr) which encodes crotonyl-CoA reductase, an enzyme that supplies 4-carbon extender units for polyketide metabolism. However, that cannot be the entire explanation, since tylosin production was restored at about 10% of the wild-type level when orf2* was re-introduced into the disrupted strain. When glycosylated precursors of tylosin were fed to the disrupted strain, they were converted to tylosin, confirming that two of the three glycosyltransferase activities associated with tylosin biosynthesis were still intact. Interestingly, however, tylactone also accumulated under such conditions and, to a much lesser extent, when tylosin was added to similar fermentations. It is concluded that glycosylated macrolides exert a pronounced positive effect on polyketide metabolism in S. fradiae.