• Microbiology and Biotechnology Letters
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• v.35 no.1
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• pp.26-29
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• 2007

Tautomycetin (TMC), which is produced by Streptomyces sp. CK4412, is a novel activated T cell-specific immunosuppressive compound with an ester bond linkage between a terminal cyclic anhydride moiety and a linear polyketide chain bearing an unusual terminal alkene. Antifungal activity against Aspergillus niger and TMC productivity assayed by HPLC using culture extracts from Streptomyces sp. CK4412 grown on solid medium adjusted at various pH were measured. The cells cultured at acidic pH (pH 4-5) medium exhibited much stronger antifungal activity as well as higher TMC productivity than those cultured at neutral pH medium, implying that the acidic pH-shock should be an efficient strategy to induce the productivity of secondary metabolites in Streptomyces culture.

• The Korean Journal of Physiology and Pharmacology
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• v.16 no.2
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• pp.145-151
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• 2012

In the present work, we studied the structure-activity relationship (SAR) of tautomycetin (TMC) and its derivatives. Further, we demonstrated the correlation between the immunosuppressive fuction, anticancer activity and protein phosphatase type 1 (PP1) inhibition of TMC and its derivatives. We have prepared some TMC derivatives via combinatorial biosynthesis, isolation from fermentation broth or chemical degradation of TMC. We found that the immunosuppressive activity was correlated with anticancer activity for TMC and its analog compounds, indicating that TMC may home at the same targets for its immunosuppressive and anticancer activities. Interestingly, TMC-F1, TMC-D1 and TMC-D2 all retained significant, albeit reduced PP1 inhibitory activity compared to TMC. However, only TMC-D2 showed immunosuppressive and anticancer activities in studies carried out in cell lines. Moreover, TMC-Chain did not show any significant inhibitory activity towards PP1 but showed strong growth inhibitory effect. This observation implicates that the maleic anhydride moiety of TMC is critical for its phosphatase inhibitory activity whereas the C1-C18 moiety of TMC is essential for the inhibition of tumor cell proliferation. Furthermore, we measured $in$ $vivo$ phosphatase activities of PP1 in MCF-7 cell extracts treated with TMC and its related compounds, and the results indicate that the cytotoxicity of TMC doesn't correlate with its $in$ $vivo$ PP1 inhibition activity. Taken together, our study suggests that the immunosuppressive and anticancer activities of TMC are not due to the inhibition of PP1. Our results provide a novel insight for the elucidation of the underlying molecular mechanisms of TMC's important biological functions.

• 한국생물공학회:학술대회논문집
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• 2005.04a
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• pp.420-422
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• 2005

TMC (Tautomycetin) is a liner polyketide immunosuppressive antifungal compound produced by Streptomyces spp. Inhibition of T cell proliferation with TMC was observed highly efficient at 100-fold lower than those needed to achieve maximal inhibition with cyclosporin A. To elucidate the biosynthetic pathway of TMC, a genomic DNA library was constructed using a E. coil-Streptomyces shuttle cosmid vector, pOJ446. The DNA libraries were screened by colony blot hybridization using several polyketide ${\beta}-ketosynthase$ (KS) probes amplified from TMC-producing Streptomyces genomic DNA using polymerase chain reaction (PCR), of which the degenerate primers were designed based on the highly conserved sequences present in KS domains of various type I polyketide synthase genes in Streptomyces species. This library construction and screening approach led to the isolation of several positive cosmid clones representing type I polyketide biosynthetic gene clusters. In addition, a Streptomyces regulatory gene called afsR2 (a global regulatory gene stimulating antibiotic production in both S. coelicolor and S. lividans) was successfully integrated into the TMC-producing Streptomyces chromosome via E. coil-Streptomyces heterologous conjugation mehtod. The more detailed results of production improvement and genetic characterization of TMC-producing Streptomyces spp. will be discussed.

• Journal of Microbiology and Biotechnology
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• v.29 no.12
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• pp.1931-1937
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• 2019

The heterologous expression of the Streptomyces natural product (NP) biosynthetic gene cluster (BGC) has become an attractive strategy for the activation, titer improvement, and refactoring of valuable and cryptic NP BGCs. Previously, a Streptomyces artificial chromosomal vector system, pSBAC, was applied successfully to the precise cloning of large-sized polyketide BGCs, including immunosuppressant tautomycetin and antibiotic pikromycin, which led to stable and comparable production in several heterologous hosts. To further validate the pSBAC system as a generally applicable heterologous expression system, the daptomycin BGC of S. roseosporus was cloned and expressed heterologously in a model Streptomyces cell factory. A 65-kb daptomycin BGC, which belongs to a non-ribosomal polypeptide synthetase (NRPS) family, was cloned precisely into the pSBAC which resulted in 28.9 mg/l of daptomycin and its derivatives in S. coelicolor M511(a daptomycin non-producing heterologous host). These results suggest that a pSBAC-driven heterologous expression strategy is an ideal approach for producing low and inconsistent Streptomyces NRPS-family NPs, such as daptomycin, which are produced low and inconsistent in native host.