Roles of
            <i>fkbN</i>
            in Positive Regulation and
            <i>tcs7</i>
            in Negative Regulation of FK506 Biosynthesis in Streptomyces sp. Strain KCTC 11604BP

Mo, SangJoon; Yoo, Young Je; Ban, Yeon Hee; Lee, Sung‐Kwon; Kim, Eunji; Suh, Joo‐Won; Yoon, Yeo Joon

doi:10.1128/aem.06766-11

Cited by 45 publications

(50 citation statements)

References 37 publications

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“…Such truncated proteins could potentially interfere with the function of intact FkbN protein, produced in the complementation experiment. All this shows, that FkbN is indispensable for FK506 production, which is in agreement with recently published results [28]. Clearly, fkbN also shows important potential for application in genetic/metabolic engineering of industrial FK506 producing strains.…”

Section: Resultssupporting

confidence: 92%

“…KCTC 11604BP has a negative regulatory role. This seems to be a somehow surprising result considering extremely high degree of similarity of both FK506 biosynthetic clusters on the level of DNA sequence [11,28]. One possible explanation is that the two strains have different general (pleiotropic) regulatory networks and/or backgrounds of primary metabolic pathways, as has been observed recently in the case of allylmalonyl-CoA extender unit biosynthesis.…”

Section: Discussionmentioning

confidence: 80%

“…In contrast to our findings, Mo et al . [28] have just recently reported that the tcs7 gene (homologue of fkbR ) from Streptomyces sp . KCTC 11604BP has a negative regulatory role.…”

Section: Discussionmentioning

confidence: 99%

“…In summary, we have clearly demonstrated, that inactivation of the fkbN gene, although completely abolishing FK506 biosynthesis, did not prevent the transcription of FK506 biosynthetic genes, contrary to the observations in Streptomyces sp . KCTC 11604BP strain, where all genes involved in biosynthesis of FK506 were silenced [28]. …”

Section: Discussionmentioning

confidence: 99%

“…In this work, we have demonstrated, that the biosynthesis of the FK506 in Streptomyces tsukubaensis NRRL 18488 is regulated by two positively-acting regulatory proteins, and remarkably, compared to the apparently closely-related strain, Streptomyces sp . KCTC 11604BP [28], it differs substantially.…”

Section: Introductionmentioning

confidence: 99%

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FK506 biosynthesis is regulated by two positive regulatory elements in Streptomyces tsukubaensis

et al. 2012

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BackgroundFK506 (Tacrolimus) is an important immunosuppressant, produced by industrial biosynthetic processes using various Streptomyces species. Considering the complex structure of FK506, it is reasonable to expect complex regulatory networks controlling its biosynthesis. Regulatory elements, present in gene clusters can have a profound influence on the final yield of target product and can play an important role in development of industrial bioprocesses.ResultsThree putative regulatory elements, namely fkbR, belonging to the LysR-type family, fkbN, a large ATP-binding regulator of the LuxR family (LAL-type) and allN, a homologue of AsnC family regulatory proteins, were identified in the FK506 gene cluster from Streptomyces tsukubaensis NRRL 18488, a progenitor of industrial strains used for production of FK506. Inactivation of fkbN caused a complete disruption of FK506 biosynthesis, while inactivation of fkbR resulted in about 80% reduction of FK506 yield. No functional role in the regulation of the FK506 gene cluster has been observed for the allN gene. Using RT-PCR and a reporter system based on a chalcone synthase rppA, we demonstrated, that in the wild type as well as in fkbN- and fkbR-inactivated strains, fkbR is transcribed in all stages of cultivation, even before the onset of FK506 production, whereas fkbN expression is initiated approximately with the initiation of FK506 production. Surprisingly, inactivation of fkbN (or fkbR) does not abolish the transcription of the genes in the FK506 gene cluster in general, but may reduce expression of some of the tested biosynthetic genes. Finally, introduction of a second copy of the fkbR or fkbN genes under the control of the strong ermE* promoter into the wild type strain resulted in 30% and 55% of yield improvement, respectively.ConclusionsOur results clearly demonstrate the positive regulatory role of fkbR and fkbN genes in FK506 biosynthesis in S. tsukubaensis NRRL 18488. We have shown that regulatory mechanisms can differ substantially from other, even apparently closely similar FK506-producing strains, reported in literature. Finally, we have demonstrated the potential of these genetically modified strains of S. tsukubaensis for improving the yield of fermentative processes for production of FK506.

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Section: Resultssupporting

confidence: 92%

Section: Discussionmentioning

confidence: 80%

“…In contrast to our findings, Mo et al . [28] have just recently reported that the tcs7 gene (homologue of fkbR ) from Streptomyces sp . KCTC 11604BP has a negative regulatory role.…”

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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FK506 biosynthesis is regulated by two positive regulatory elements in Streptomyces tsukubaensis

et al. 2012

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Immunosuppressants: Remarkable Microbial Products

Vaishnav¹,

Yoo

Yoon

et al. 2014

Bioactive Natural Products

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Generation of Streptomyces hygroscopicus cell factories with enhanced ascomycin production by combined elicitation and pathway‐engineering strategies

Wang

Yuan

et al. 2019

Biotech & Bioengineering

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Ascomycin (FK520) is a macrocyclic antibiotic that also exhibits antifungal and immunosuppressive activity. However, its relatively low titer and yield have hampered commercial application. Here, we have successfully constructed an efficient ascomycinproducing strain of Streptomyces hygroscopicus with high titer and yield, using a novel combinatorial engineering approach based on the identification of targets involved in both metabolic and transcriptional regulation. First, we investigated the effects of different chemicals on ascomycin accumulation and found that dimethyl sulfoxide best stimulated ascomycin overproduction. We next compared intracellular metabolic and transcriptional profiles after dimethyl sulfoxide and control treatments and identified potential target genes (zwf and aroA, involved in metabolic precursor pathways; and luxR, iclR, fadR, and fkbN, involved in transcriptional regulation). These candidate genes were then engineered to produce strains with individual and combinatorial overexpression. Combined overexpression of aroA, fkbN, and luxR resulted in the highest yield of ascomycin (1258.30 ± 33.49 mg/L), 4.12-fold higher than the control yield (305.60 ± 16.90 mg/L). This integrative multilevel approach identified novel determinants involved in both metabolic and transcriptional regulation, resulting in the diversion of carbon flux towards ascomycin accumulation. This approach could be applied to boost the production of a variety of useful bacterial metabolites.

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Roles of fkbN in Positive Regulation and tcs7 in Negative Regulation of FK506 Biosynthesis in Streptomyces sp. Strain KCTC 11604BP

Cited by 45 publications

References 37 publications

FK506 biosynthesis is regulated by two positive regulatory elements in Streptomyces tsukubaensis

FK506 biosynthesis is regulated by two positive regulatory elements in Streptomyces tsukubaensis

Immunosuppressants: Remarkable Microbial Products

Generation of Streptomyces hygroscopicus cell factories with enhanced ascomycin production by combined elicitation and pathway‐engineering strategies

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