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. 2009 Mar 15;17(6):2147-53.
doi: 10.1016/j.bmc.2008.10.074. Epub 2008 Nov 5.

Engineered production of iso-migrastatin in heterologous Streptomyces hosts

Zhiyang Feng  1 , Liyan Wang, Scott R Rajski, Zhinan Xu, Marie F Coeffet-LeGal, Ben Shen
Affiliations

Engineered production of iso-migrastatin in heterologous Streptomyces hosts

Zhiyang Feng et al. Bioorg Med Chem. .

Abstract

Glutarimide-containing polyketides such as migrastatin (MGS) are well known for their ability to inhibit tumor cell migration. We have previously shown that MGS is derived from iso-migrastatin (iso-MGS) via a H(2)O-mediated ring-expansion rearrangement. A bacterial artificial chromosome (BAC) library of Streptomyces platensis NRRL18993, an iso-MGS producer, was constructed. From this library, pBS11001, a BAC clone harboring the intact iso-MGS biosynthetic gene cluster, was identified. Mobilization of pBS11001 into five heterologous Streptomyces hosts afforded recombinant strains, SB11001, SB11002, SB11003, SB11004, and SB11005, respectively. Under a standard set of media and fermentation conditions, the recombinant strains all produced the same profile of iso-MGS as that of S. platensis NRRL18993. These findings highlight the strength and flexibility of the BAC-based technology for natural product production and engineering in heterologous Streptomyces model hosts.

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Figures

Figure 1
Figure 1
Glutarimide-containing polyketides isolated from S. platensis NRRL18993: iso-migrastatin and its H2O-mediated ring-expansion and ring-opening rearrangements to migrastatin and dorrigocins.
Figure 2
Figure 2
BAC-based technology for natural product production and engineering in heterologous Streptomyces hosts: (A) the pStreptoBAC V vector, featuring two BamHI sites, the apramycin resistance marker (AmR), oriT, and ΦC31 and (B) the pBS11001 clone with 65-kb DNA insert from S. platensis NRRL18993 that harbors the intact mgs biosynthetic gene cluster.
Figure 3
Figure 3
Analysis of BAC clones of S. platensis NRRL18992 library. Eighteen randomly picked BAC clones were digested with HindIII and the digested DNAs were analyzed by pulsed field gel electrophoresis (PFGE). Lanes 1 to 18, HindIII-digested BAC DNAs; M, DNA marker, with relevant sizes noted on right.
Figure 4
Figure 4
HPLC analysis of extracts from recombinant strains SB11001 (II), SB11002 (III), SB11003 (IV), SB11004 (V), SB11005 (VI), with S. platensis NRRL18993 as a control (I), cultured in (A) B2 or (B) R2YE medium. ▼, iso-MGS. Normalization of iso-MGS signal intensity between panel A and B analyses required that five times as much R2YE-derived extract be injected compared to the corresponding B2-derived extract (see experimental section 5.5).
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References

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