Acyldepsipeptide HDAC inhibitor production induced in Burkholderia thailandensis

Abstract

Natural product gene clusters are often tightly regulated, resulting in gene cluster silencing in laboratory fermentation studies. The systematic overexpression of transcription factors (TFs) associated with biosynthetic gene clusters found in the genome of Burkholderia thailandensis E264 identified a set of TFs that, when overexpressed, alter the secondary metabolome of this bacterium. The isolation and characterization of burkholdacs A and B, two new acyldepsitripeptide histone deacetylase inhibitors produced by B. thailandensis overexpressing the TF bhcM, is reported.

Figure 1

Activating cryptic metabolite production via transcription factor expression: (A) Transcription factors putatively associated with B. thailandensis E264 biosynthetic gene clusters were PCR amplified, individually cloned downstream of a P_tac promoter and the resulting constructs were then transformed back into B. thailandensis E264. Culture broth extracts from E264 strains over-expressing each transcription factor were assessed for the presence of metabolites not seen in extracts from similarly treated vector control cultures. (B) HPLC analysis of the ethyl acetate extract from cultures of E264 overexpressing TF Bth_I2369. Peaks marked 1 and 2 on this trace correspond to burkholdacs A and B, respectively (Figure 2). (C) The predicted NRPS/PKS gene cluster directly adjacent to TF Bth_I2369 is shown (blue, predicted NRPS/PKS genes; red, cloned transcription factor).

Figure 2

NMR assignments of the burholdac core structure.

Figure 3

Natural depsipeptides related to burkholdacs A and B.

Figure 4

(a) DEP and BHC gene clusters. (b) The late stages of burkholdac biosynthesis (bhcDEK) are unique to the BHC cluster. ACP/PCP, acyl or peptidyl carrier protein; C, condensation; A, adenylation; E, epimerization; KS, ketosynthase; KR, ketoreductase; TE, thioesterase. Superscript “nf” notes nonfunctional domain. Gene and domain functional predictions appear in Table S2.