An unexpected interaction between the modular polyketide synthases, erythromycin DEBS1 and pikromycin PikAIV, leads to efficient triketide lactone synthesis

Beom Seok Kim, T. Ashton Cropp, Galina Florova, Yuko Lindsay, David H. Sherman, Kevin A. Reynolds

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

An unusual feature of the 6-module pikromycin polyketide synthase (PikPKS, PikAI-PikAIV) of S. venezuelae is the ability to generate both 12- and 14-membered ring macrolides. The PikAIV component containing the last extension module and a thioesterase domain is responsible for generating both of these products. In the case of the 12-membered ring macrolide, an acyl-enzyme intermediate on PikAIII is able to efficiently "skip" the last extension step and is cyclized by the TE domain of PikAIV, presumably as a result of a PikAIII-PikAIV interaction. Herein we report that plasmid-based expression (pBK3) of DEBS1, which comprises the loading domain and the first two modules of the Saccharopolyspora erythrea 6-deoxyerythronolide B synthase, in S. venezuelae leads to efficient 15 ± 3 mg/L production of triketide lactone products (TKLs). Comparable levels of TKLs were observed with a plasmid (pBK1) which expressed DEBS1 fused to a TE domain (DEBS1-TE). These results are in stark contrast to previous in vivo and in vitro analyses, where only DEBS1-TE efficiently produces TKLs. Levels of TKLs decreased dramatically with expression of DEBS1 in both pikAIV and pikAIII-pikAIV deletion hosts (0.5 mg/L), but not DEBS1-TE, and could be partially restored by addition of a PikAIV complementation plasmid. These data suggest that PikAIV is able to efficiently catalyze formation of 6-membered lactone ring products from acyl-bound intermediates on DEBS1 in a manner analogous to that observed for 12-membered macrolide products from PikAIII. Significant sequence similarity and length of the C-terminal linker region of PikAIII and DEBS1 suggest that this region may be responsible for the interaction with PikAIV. A replacement of this linker region of DEBS1 with the corresponding region of PikAI led to a 95% decrease in TKL levels in S. venezuelae, consistent with this hypothesis.

Original languageEnglish
Pages (from-to)10827-10833
Number of pages7
JournalBiochemistry
Volume41
Issue number35
DOIs
Publication statusPublished - 2002 Sep 3
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry

Fingerprint Dive into the research topics of 'An unexpected interaction between the modular polyketide synthases, erythromycin DEBS1 and pikromycin PikAIV, leads to efficient triketide lactone synthesis'. Together they form a unique fingerprint.

  • Cite this