Overexpression of specific proton motive force-dependent transporters facilitate the export of surfactin in Bacillus subtilis

Li, Xu; Yang, Huan; Zhang, Donglai; Li, Xue; Yu, Huimin; Shen, Zhongyao

doi:10.1007/s10295-014-1527-z

Cited by 56 publications

(46 citation statements)

References 48 publications

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“…The wild surfactin producer, B. subtilis THY‐7, was isolated from a soil sample and deposited at the China General Microbiological Culture Collection Center (CGMCC No. 8906) (Li et al, ). E. coli TOP10 were used for gene cloning and plasmid construction, and E. coli BL21 were used for lacZ reporter gene cloning.…”

Section: Methodsmentioning

confidence: 99%

“…Surfactin concentrations were determined by high performance liquid chromatography (HPLC) using an LC‐20A HPLC system (Shimadzu, Kyoto, Japan) equipped with an ODS‐SP column (250 × 4.6 mm, 5 μm; GL Sciences, Kyoto, Japan) and a DA detector (Shimadzu) as previously described by Li et al (). Surfactin standards were purchased from Sigma (Shanghai, China), and experiments were performed in triplicate.…”

Section: Methodsmentioning

confidence: 99%

“…Surfactin is natively synthesized by surfactin synthase (SrfA), a seven module non‐ribosomal peptide synthase (NRPS), and then transported outside the cells during fermentation (Roongsawang et al, ; Tapi et al, ). In light of this mechanism, previous studies have applied multiple genetic strategies to various B. subtili s species, including promoter replacement (Bechet et al, ; Fickers et al, ; Sun et al, ), and overexpression of specific extracellular signaling peptides (Jung et al, ), assistant proteins and surfactin transporters (Li et al, ; Sekhon et al, ). The maximum surfactin titers achieved through these approaches were 3.87, 0.14, and 2.13 g/L, respectively.…”

Section: Introductionmentioning

confidence: 99%

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In situ enhancement of surfactin biosynthesis in Bacillus subtilis using novel artificial inducible promoters

Song

et al. 2016

Biotech & Bioengineering

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Surfactin-family lipopeptides are green biosurfactants with substantial industrial potential. The major problem prohibiting surfactin use is the low titer of the wild producer, Bacillus subtilis. Using transcriptomic analysis, four strong promoters, PgroE, Pcdd, PrplK, and PsspE, were identified and cloned from the genome of B. subtilis THY-7, a novel surfactin producer that has been identified from soil with a 0.55 g/L surfactin titer. An optimal promoter, PgroE, was selected to replace the native THY-7 surfactin synthase (SrfA) promoter through single-cross homologous recombination; however, the resulting engineered strain containing the PgroE substitution did not synthesize surfactin. The sucrose-inducible promoters PsacB and PsacP were then substituted in place of PsrfA, and the resulting engineered strains produced 1.09 and 0.22 g/L surfactin, respectively. An artificial, sucrose-inducible Pg1 promoter was produced through fusion of the PgroE and PsacB ribonucleic antiterminator (RAT), and the engineered strain containing the Pg1-substitution produced a surfactin titer of 1.44 g/L. An artificial IPTG-inducible promoter, Pg2, was constructed from a PgroE-lacO fusion and then substituted for the chromosomal PsrfA locus, and the surfactin titer of the resulting THY-7/Pg2-srfA increased to 5.98 g/L. The driving capacity of Pg2 was further improved by the inclusion of two point mutations in the -35 and -10 regions to produce the novel promoter Pg3. Pg3 exhibited super-strong activity as measured by lacZ reporter gene overexpression (approximately 3000 U). The Pg3-substitution strain THY-7/Pg3-srfA produced up to 9.74 g/L surfactin in a 5 L fermentor. The maximum productivity was 0.30 g/L/h, and the greatest yield reached 0.14 g surfactin/g sucrose. Biotechnol. Bioeng. 2017;114: 832-842. © 2016 Wiley Periodicals, Inc.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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In situ enhancement of surfactin biosynthesis in Bacillus subtilis using novel artificial inducible promoters

Song

et al. 2016

Biotech & Bioengineering

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“…cereus group strains, but a recent study demonstrated that an orthologous pump is involved in the efflux of a surfactin in B . subtilis [37]. Surfactin has been shown by a number of studies to be essential for formation of mature biofilms by B .…”

Section: Resultsmentioning

confidence: 99%

“…subtilis (Table 4). Functional analyses of YerP recently demonstrated that overexpression of this pump in its native host resulted in increased secretion of endogenously produced surfactin into the supernatant [37]. YerP is also known to be involved in surfactin resistance in strains that do not produce an endogenous surfactin and can mediate resistance to acriflavine and ethidium [50].…”

Section: Resultsmentioning

confidence: 99%

The putative drug efflux systems of the Bacillus cereus group

et al. 2017

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The Bacillus cereus group of bacteria includes seven closely related species, three of which, B. anthracis, B. cereus and B. thuringiensis, are pathogens of humans, animals and/or insects. Preliminary investigations into the transport capabilities of different bacterial lineages suggested that genes encoding putative efflux systems were unusually abundant in the B. cereus group compared to other bacteria. To explore the drug efflux potential of the B. cereus group all putative efflux systems were identified in the genomes of prototypical strains of B. cereus, B. anthracis and B. thuringiensis using our Transporter Automated Annotation Pipeline. More than 90 putative drug efflux systems were found within each of these strains, accounting for up to 2.7% of their protein coding potential. Comparative analyses demonstrated that the efflux systems are highly conserved between these species; 70–80% of the putative efflux pumps were shared between all three strains studied. Furthermore, 82% of the putative efflux system proteins encoded by the prototypical B. cereus strain ATCC 14579 (type strain) were found to be conserved in at least 80% of 169 B. cereus group strains that have high quality genome sequences available. However, only a handful of these efflux pumps have been functionally characterized. Deletion of individual efflux pump genes from B. cereus typically had little impact to drug resistance phenotypes or the general fitness of the strains, possibly because of the large numbers of alternative efflux systems that may have overlapping substrate specificities. Therefore, to gain insight into the possible transport functions of efflux systems in B. cereus, we undertook large-scale qRT-PCR analyses of efflux pump gene expression following drug shocks and other stress treatments. Clustering of gene expression changes identified several groups of similarly regulated systems that may have overlapping drug resistance functions. In this article we review current knowledge of the small molecule efflux pumps encoded by the B. cereus group and suggest the likely functions of numerous uncharacterised pumps.

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Genetic engineering of the branched‐chain fatty acid biosynthesis pathway to enhance surfactin production from Bacillus subtilis

Jing

Wei

Liu

et al. 2022

Biotech and App Biochem

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Surfactin, which is composed of a β‐hydroxy fatty acid chain and a peptide ring, has drawn considerable attention due to its potential applications in the biomedicine, bioremediation, and petroleum industries. However, the low yield of surfactin from wild strains still restricts its industrial applications. In this study, eight genes relevant to the fatty acid biosynthesis pathway were targeted to enhance surfactin production, and high surfactin‐yielding strains with potential industrial applications were obtained. When ldeHA and acc were co‐overexpressed, the surfactin yield of recombinant strains TDS8 and TPS8 increased to 1.55‐ and 1.19‐fold of their parental strains, respectively, again proving that the conversion of acetyl‐coenzyme A (CoA) to malonyl‐CoA is the rate‐limiting step in fatty acid biosynthesis. Furthermore, changes in surfactin isoforms of recombinant strain TPS8 suggest that the fatty acid precursor synthesis pathway can be modified to improve the proportion of different isoforms. In addition, the deletion of lpdV, which is responsible for the conversion of α‐ketoacyl‐CoA precursors, resulted in a sharp decrease in surfactin production, further demonstrating the importance of branched‐chain fatty acid biosynthesis in surfactin production. This work will facilitate the design and construction of more efficiently engineered strains for surfactin production and further extend industrial applications.

show abstract

Overexpression of specific proton motive force-dependent transporters facilitate the export of surfactin in Bacillus subtilis

Cited by 56 publications

References 48 publications

In situ enhancement of surfactin biosynthesis in Bacillus subtilis using novel artificial inducible promoters

In situ enhancement of surfactin biosynthesis in Bacillus subtilis using novel artificial inducible promoters

The putative drug efflux systems of the Bacillus cereus group

Genetic engineering of the branched‐chain fatty acid biosynthesis pathway to enhance surfactin production from Bacillus subtilis

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