Dataset Information

Characterization of SAV7471, a TetR-family transcriptional regulator involved in the regulation of coenzyme A metabolism in Streptomyces avermitilis.

ABSTRACT: The role of a tetR transcriptional regulatory gene (SAV7471) in avermectin production in the Gram-positive soil bacterium Streptomyces avermitilis was investigated by gene deletion, complementation, and overexpression experiments. Gene deletion of the SAV7471 open reading frame resulted in avermectin overproduction. The deletion also resulted in overexpression of SAV7472-SAV7473 transcripts, which encode a protein of unknown function and a flavoprotein possibly involved in pantothenate and coenzyme A (CoA) metabolism. EMSAs and footprinting assays showed that SAV7471 can bind to two palindromic sequences with high similarity in the intergenic region between SAV7471 and SAV7472, a region that contains the apparent transcription start sites for each gene detected by rapid amplification of 5' cDNA ends (5'-RACE). In addition to SAV7472-SAV7473, at least two genes (SAV1104 and SAV1258) involved in CoA metabolism are negatively controlled by SAV7471. By negatively regulating the transcription of the target genes SAV7472-SAV7473 and other genes involved in CoA metabolism, SAV7471 may affect cellular metabolic flux and may thereby indirectly regulate avermectin biosynthesis.

SUBMITTER: Liu Y

PROVIDER: S-EPMC3807474 | biostudies-literature | 2013 Oct

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Characterization of SAV7471, a TetR-family transcriptional regulator involved in the regulation of coenzyme A metabolism in Streptomyces avermitilis.

Liu Yanping Y Yan Tingting T Jiang Libin L Wen Ying Y Song Yuan Y Chen Zhi Z Li Jilun J

Journal of bacteriology 20130726 19

The role of a tetR transcriptional regulatory gene (SAV7471) in avermectin production in the Gram-positive soil bacterium Streptomyces avermitilis was investigated by gene deletion, complementation, and overexpression experiments. Gene deletion of the SAV7471 open reading frame resulted in avermectin overproduction. The deletion also resulted in overexpression of SAV7472-SAV7473 transcripts, which encode a protein of unknown function and a flavoprotein possibly involved in pantothenate and coenz ...[more]

PMID: 23893108

Similar Datasets

Project description:Malonyl coenzyme A (malonyl-CoA) and methylmalonyl-CoA are the most common extender units for the biosynthesis of fatty acids and polyketides in Streptomyces, an industrially important producer of polyketides. Carboxylation of acetyl- and propionyl-CoAs is an essential source of malonyl- and methylmalonyl-CoAs; therefore, acyl-CoA carboxylases (ACCases) play key roles in primary and secondary metabolism. The regulation of the expression of ACCases in Streptomyces spp. has not been investigated previously. We characterized a TetR family transcriptional repressor, AccR, that mediates intracellular acetyl-, propionyl-, methylcrotonyl-, malonyl-, and methylmalonyl-CoA levels by controlling the transcription of genes that encode the main ACCase and enzymes associated with branched-chain amino acid metabolism in S. avermitilis AccR bound to a 16-nucleotide palindromic binding motif (GTTAA-N6-TTAAC) in promoter regions and repressed the transcription of the accD1A1-hmgL-fadE4 operon, echA8, echA9, and fadE2, which are involved in the production and assimilation of acetyl- and propionyl-CoAs. Methylcrotonyl-, propionyl-, and acetyl-CoAs acted as effectors to release AccR from its target DNA, resulting in enhanced transcription of target genes by derepression. The affinity of methylcrotonyl- and propionyl-CoAs to AccR was stronger than that of acetyl-CoA. Deletion of accR resulted in increased concentrations of short-chain acyl-CoAs (acetyl-, propionyl-, malonyl-, and methylmalonyl-CoAs), leading to enhanced avermectin production. Avermectin production was increased by 14.5% in an accR deletion mutant of the industrial high-yield strain S. avermitilis A8. Our findings clarify the regulatory mechanisms that maintain the homeostasis of short-chain acyl-CoAs in Streptomyces IMPORTANCE Acyl-CoA carboxylases play key roles in primary and secondary metabolism. However, the regulation of ACCase genes transcription in Streptomyces spp. remains unclear. Here, we demonstrated that AccR responded to intracellular acetyl-, propionyl-, and methylcrotonyl-CoA availability and mediated transcription of the genes related to production and assimilation of these compounds in S. avermitilis When intracellular concentrations of these compounds are low, AccR binds to target genes and represses their transcription, resulting in low production of malonyl- and methylmalonyl-CoAs. When intracellular acetyl-, propionyl-, and methylcrotonyl-CoA concentrations are high, these compounds bind to AccR to dissociate AccR from target DNA, promoting the conversion of these compounds to malonyl- and methylmalonyl-CoAs. This investigation revealed how AccR coordinates short-chain acyl-CoA homeostasis in Streptomyces.

Project description:Propanol stimulates erythromycin biosynthesis by increasing the supply of propionyl coenzyme A (propionyl-CoA), a starter unit of erythromycin production in Saccharopolyspora erythraea Propionyl-CoA is assimilated via propionyl-CoA carboxylase to methylmalonyl-CoA, an extender unit of erythromycin. We found that the addition of n-propanol or propionate caused a 4- to 16-fold increase in the transcriptional levels of the SACE_3398-3400 locus encoding propionyl-CoA carboxylase, a key enzyme in propionate metabolism. The regulator PccD was proved to be directly involved in the transcription regulation of the SACE_3398-3400 locus by EMSA and DNase I footprint analysis. The transcriptional levels of SACE_3398-3400 were upregulated 15- to 37-fold in the pccD gene deletion strain (?pccD) and downregulated 3-fold in the pccD overexpression strain (WT/pIB-pccD), indicating that PccD was a negative transcriptional regulator of SACE_3398-3400. The ?pccD strain has a higher growth rate than that of the wild-type strain (WT) on Evans medium with propionate as the sole carbon source, whereas the growth of the WT/pIB-pccD strain was repressed. As a possible metabolite of propionate metabolism, methylmalonic acid was identified as an effector molecule of PccD and repressed its regulatory activity. A higher level of erythromycin in the ?pccD strain was observed compared with that in the wild-type strain. Our study reveals a regulatory mechanism in propionate metabolism and suggests new possibilities for designing metabolic engineering to increase erythromycin yield.IMPORTANCE Our work has identified the novel regulator PccD that controls the expression of the gene for propionyl-CoA carboxylase, a key enzyme in propionyl-CoA assimilation in S. erythraea PccD represses the generation of methylmalonyl-CoA through carboxylation of propionyl-CoA and reveals an effect on biosynthesis of erythromycin. This finding provides novel insight into propionyl-CoA assimilation, and extends our understanding of the regulatory mechanisms underlying the biosynthesis of erythromycin.

Dataset Information

Characterization of SAV7471, a TetR-family transcriptional regulator involved in the regulation of coenzyme A metabolism in Streptomyces avermitilis.

Publications

Characterization of SAV7471, a TetR-family transcriptional regulator involved in the regulation of coenzyme A metabolism in Streptomyces avermitilis.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets