New erythromycin derivatives from Saccharopolyspora erythraea using sugar O‐methyltransferases from the spinosyn biosynthetic gene cluster

Abstract

SummaryUsing a previously developed expression system based on the erythromycin-producing strain of Saccharopolyspora erythraea, O-methyltransferases from the spinosyn biosynthetic gene cluster of Saccharopolyspora spinosa have been shown to modify a rhamnosyl sugar attached to a 14-membered polyketide macrolactone. The spnI, spnK and spnH methyltransferase genes were expressed individually in the S. erythraea mutant SGT2, which is blocked both in endogenous macrolide biosynthesis and in ery glycosyltransferases eryBV and eryCIII. Exogenous 3-O-rhamnosyl-erythronolide B was efficiently converted into 3-O-(2 0 -O-methylrhamnosyl)-erythronolide B by the S. erythraea SGT2 (spnI ) strain only.was, in turn, fed to a culture of S. erythraea SGT2 (spnK ), 3-O-(2 0 ,3 0 -bis-O-methylrhamnosyl)-erythronolide B was identified in the culture supernatant, whereas S. erythraea SGT2 (spnH ) was without effect. These results confirm the identity of the 2 0 -and 3 0 -Omethyltransferases, and the specific sequence in which they act, and they demonstrate that these methyltransferases may be used to methylate rhamnose units in other polyketide natural products with the same specificity as in the spinosyn pathway. In contrast, 3-O-(2 0 ,3 0 -bis-O-methylrhamnosyl)-erythronolide B was found not to be a substrate for the 4 0 -O-methyltransferase SpnH. Although rhamnosylerythromycins did not serve directly as substrates for the spinosyn methyltransferases, methylrhamnosylerythromycins were obtained by subsequent conversion of the corresponding methylrhamnosyl-erythronolide precursors using the S. erythraea strain SGT2 housing EryCIII, the desosaminyltransferase of the erythromycin pathway. 3-O-(2 0 -O-methylrhamnosyl)-erythromycin D was tested and found to be significantly active against a strain of erythromycin-sensitive Bacillus subtilis.

Natural‐Product Sugar Biosynthesis and Enzymatic Glycodiversification

“…Finally, the individual expression of several l-rhamnosyl-O-methyltransferases from the spinosyn biosynthetic pathway of Saccharopolyspora spinosa in the SGT2 triple mutant demonstrated that two of these methyltransferases (SpnI and SpnK) could sequentially methylate the oxygen atoms of the 2'-and 3'-OH groups, respectively, of exogenously fed 3-rhamnosyl erythronolide B (214) to give 217 and 218 (Scheme 17, path D). [263] (43) is the sugar donor used for methymycin and pikromycin (219-221, 187) biosynthesis in Streptomyces venezuelae. As shown in Scheme 18, disruption of the dimethyltransferase gene desVI resulted in the accumulation of macrolide analogues carrying 3-N-acetylamino-3,4,6-trideoxy-d-glucose (222) in place of d-desosamine.…”

Engineered biosynthesis of hybrid macrolide polyketides containing d-angolosamine and d-mycaminose moieties

“…1). [5][6][7][8] Here we report the successful use of biosynthetic gene cassettes for the heterologous production and transfer of several deoxyhexoses to polyketide aglycones in place of the natural substituents. These are changes of potential relevance for generating compounds with altered and desirable activities.…”

Molecular cloning, sequence analysis and functional characterization of the gene cluster for biosynthesis of K-252a and its analogs

“…On the other hand, another methyltransferase encoded by nokM may possibly be used for the biosynthesis of, yet unidentified, O-methylated K-252d (most likely 2 0 -or 3 0 -O-methyl-K-252d) in the same strain, as nokM was found to share higher sequence similarity with spnI (a 2 0 -or 3 0 -O-methyltransferase gene) than spnK and spnH in methylation of spinosyn rhamnose. 34,52 Based on our findings here and in the companion paper, we may thus postulate a more complete biosynthetic pathway, as illustrated in Scheme 1, to account for biosyntheses of indolocarbazole metabolites in Nocardiopsis sp. K-252.…”