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Haruo Ikeda - One of the best experts on this subject based on the ideXlab platform.

  • Hydroxylation of Compactin (ML-236B) by CYP105D7 (SAV_7469) from Streptomyces avermitilis.
    Journal of microbiology and biotechnology, 2017
    Co-Authors: Qiuping Yao, Haruo Ikeda, Ling Liu, Shinya Fushinobu
    Abstract:

    Compactin and pravastatin are competitive cholesterol biosynthesis inhibitors of 3-hydroxy-3-methylglutaryl-CoA reductase and belong to the statin drugs; however, the latter shows superior pharmacokinetic characteristics. Previously, we reported that the bacterial P450, CYP105D7, from Streptomyces avermitilis can catalyze the hydroxylation of 1-deoxypentalenic acid, diclofenac, and naringenin. Here, we demonstrate that CYP105D7 could also catalyze compactin hydroxylation in vitro. In the presence of both bacterial and cyanobacterial redox partner systems with an NADPH regeneration system, the reaction produced two hydroxylated products, including pravastatin (hydroxylated at the C6 position). The steady-state kinetic parameters were measured using the redox partners of putidaredoxin and its reductase. The Km and kcat values for compactin were 39.1 ± 8.8 µM and 1.12 ± 0.09 min-1, respectively. The kcat/Km value for compactin (0.029 min-1·µM-1) was lower than that for diclofenac (0.114 min-1·µM-1). Spectroscopic analysis showed that CYP105D7 binds to compactin with a Kd value of 17.5 ± 3.6 µM. Molecular docking analysis was performed to build a possible binding model of compactin. Comparisons of different substrates with CYP105D7 were conclusively illustrated for the first time.

  • Hydroxylation of flavanones by cytochrome P450 105D7 from Streptomyces avermitilis
    Journal of Molecular Catalysis B: Enzymatic, 2016
    Co-Authors: Ling Liu, Haruo Ikeda, Qiuping Yao, Shinya Fushinobu
    Abstract:

    Flavanones have a wide range of pharmacological activities. Previously, we showed that CYP105D7, a cytochrome P450, from Streptomyces avermitilis can catalyze hydroxylation of diclofenac at the C4′ position. Here, we demonstrated that CYP105D7 also catalyzes hydroxylation of two flavanones, naringenin and pinocembrin. Naringenin was hydroxylated at the 3′-position in a regiospecific manner to yield eriodictyol. Spectroscopic analyses showed that CYP105D7 binds to naringenin and pinocembrin in a weakly cooperative manner with an affinity of 103 μM and 52 μM, and a Hill coefficient of 1.25 and 1.47, respectively. A possible binding model of naringenin was investigated by molecular-docking analyses. The substrate-binding pocket of CYP105D7 is sufficiently wide to accommodate two naringenin molecules simultaneously, and the C3′ atom of the proximal molecule is in the appropriate location for aromatic hydroxylation.

  • Total synthesis and absolute configuration of avenolide, extracellular factor in Streptomyces avermitilis
    The Journal of Antibiotics, 2011
    Co-Authors: Miho Uchida, Haruo Ikeda, Satoshi Takamatsu, Shiho Arima, Kiyoko T Miyamoto, Shigeru Kitani, Takuya Nihira, Tohru Nagamitsu
    Abstract:

    The first total synthesis of extracellular factor, “Avenolide”, in Streptomyces avermitilis has been achieved using a convergent approach. The stereogenic centers in two key segments were installed using Sharpless epoxidation and dihydroxylation. This synthetic study allowed the determination of the absolute configuration of avenolide as 4 S ,10 R , and yielded important information on its structure–activity relationship.

  • avenolide a Streptomyces hormone controlling antibiotic production in Streptomyces avermitilis
    Proceedings of the National Academy of Sciences of the United States of America, 2011
    Co-Authors: Shigeru Kitani, Satoshi Omura, Miho Uchida, Satoshi Takamatsu, Kiyoko T Miyamoto, Tohru Nagamitsu, Elisa Herawati, Hiroyuki Iguchi, Kouhei Nishitomi, Haruo Ikeda
    Abstract:

    Gram-positive bacteria of the genus Streptomyces are industrially important microorganisms, producing >70% of commercially important antibiotics. The production of these compounds is often regulated by low-molecular-weight bacterial hormones called autoregulators. Although 60% of Streptomyces strains may use γ-butyrolactone–type molecules as autoregulators and some use furan-type molecules, little is known about the signaling molecules used to regulate antibiotic production in many other members of this genus. Here, we purified a signaling molecule (avenolide) from Streptomyces avermitilis—the producer of the important anthelmintic agent avermectin with annual world sales of $850 million—and determined its structure, including stereochemistry, by spectroscopic analysis and chemical synthesis as (4S,10R)-10-hydroxy-10-methyl-9-oxo-dodec-2-en-1,4-olide, a class of Streptomyces autoregulator. Avenolide is essential for eliciting avermectin production and is effective at nanomolar concentrations with a minimum effective concentration of 4 nM. The aco gene of S. avermitilis, which encodes an acyl-CoA oxidase, is required for avenolide biosynthesis, and homologs are also present in Streptomyces fradiae, Streptomyces ghanaensis, and Streptomyces griseoauranticus, suggesting that butenolide-type autoregulators may represent a widespread and another class of Streptomyces autoregulator involved in regulating antibiotic production.

  • characterization of a silent sesquiterpenoid biosynthetic pathway in Streptomyces avermitilis controlling epi isozizaene albaflavenone biosynthesis and isolation of a new oxidized epi isozizaene metabolite
    Microbial Biotechnology, 2011
    Co-Authors: Satoshi Takamatsu, David E Cane, Mamoru Komatsu, Xin Lin, Ayako Nara, Haruo Ikeda
    Abstract:

    The genome‐sequenced, Gram‐positive bacterium Streptomyces avermitilis harbours an orthologue (SAV_3032) of the previously identified epi‐isozizaene synthase (SCO5222) in Streptomyces coelicolor A3(2). The sav3032 is translationally coupled with the downstream sav3031 gene encoding the cytochrome P450 CYP170A2 analogous to SCO5223 (CYP170A1) of S. coelicolor A3(2), which exhibits a similar translation coupling. Streptomyces avermitilis did not produce epi‐isozizaene or any of its oxidized derivatives, albaflavenols and albaflavenone, under in any culture conditions examined. Nonetheless, recombinant SAV_3032 protein expressed in Escherichia coli catalysed the Mg2+‐dependent cyclization of farnesyl diphosphate to epi‐isozizaene. To effect the production of epi‐isozizaene in S. avermitilis, the sav3032 gene was cloned and placed under control of a copy of the native S. avermitilis promoter rpsJp (sav4925). The derived expression construct was introduced by transformation into a large‐deletion mutant of S. avermitilis SUKA16 and the resulting transformants accumulated epi‐isozizaene. The previously characterized oxidized epi‐isozizaene metabolites (4R)‐ and (4S)‐albaflavenols and albaflavenone, as well as a previously undescribed doubly oxidized epi‐isozizaene derivative were isolated from cultures of S. avermitilis SUKA16 transformants in which sav3032 was coexpressed with the P450‐encoding sav3031. This new metabolite was identified as 4β,5β‐epoxy‐2‐epi‐zizaan‐6β‐ol which is most likely formed by oxidation of (4S)‐albaflavenol.

Satoshi Omura - One of the best experts on this subject based on the ideXlab platform.

  • engineered Streptomyces avermitilis host for heterologous expression of biosynthetic gene cluster for secondary metabolites
    ACS Synthetic Biology, 2013
    Co-Authors: Mamoru Komatsu, Satoshi Omura, Kyoko Komatsu, Hanae Koiwai, Yuuki Yamada, Ikuko Kozone, Miho Izumikawa, Junko Hashimoto, Motoki Takagi, Kazuo Shinya
    Abstract:

    An industrial microorganism, Streptomyces avermitilis, which is a producer of anthelmintic macrocyclic lactones, avermectins, has been constructed as a versatile model host for heterologous expression of genes encoding secondary metabolite biosynthesis. Twenty of the entire biosynthetic gene clusters for secondary metabolites were successively cloned and introduced into a versatile model host S. avermitilis SUKA17 or 22. Almost all S. avermitilis transformants carrying the entire gene cluster produced metabolites as a result of the expression of biosynthetic gene clusters introduced. A few transformants were unable to produce metabolites, but their production was restored by the expression of biosynthetic genes using an alternative promoter or the expression of a regulatory gene in the gene cluster that controls the expression of biosynthetic genes in the cluster using an alternative promoter. Production of metabolites in some transformants of the versatile host was higher than that of the original produc...

  • Engineered Streptomyces avermitilis Host for Heterologous Expression of Biosynthetic Gene Cluster for Secondary Metabolites
    2013
    Co-Authors: Mamoru Komatsu, Satoshi Omura, Kyoko Komatsu, Hanae Koiwai, Yuuki Yamada, Ikuko Kozone, Miho Izumikawa, Junko Hashimoto, Motoki Takagi, Kazuo Shin-ya
    Abstract:

    An industrial microorganism, Streptomyces avermitilis, which is a producer of anthelmintic macrocyclic lactones, avermectins, has been constructed as a versatile model host for heterologous expression of genes encoding secondary metabolite biosynthesis. Twenty of the entire biosynthetic gene clusters for secondary metabolites were successively cloned and introduced into a versatile model host S. avermitilis SUKA17 or 22. Almost all S. avermitilis transformants carrying the entire gene cluster produced metabolites as a result of the expression of biosynthetic gene clusters introduced. A few transformants were unable to produce metabolites, but their production was restored by the expression of biosynthetic genes using an alternative promoter or the expression of a regulatory gene in the gene cluster that controls the expression of biosynthetic genes in the cluster using an alternative promoter. Production of metabolites in some transformants of the versatile host was higher than that of the original producers, and cryptic biosynthetic gene clusters in the original producer were also expressed in a versatile host

  • avenolide a Streptomyces hormone controlling antibiotic production in Streptomyces avermitilis
    Proceedings of the National Academy of Sciences of the United States of America, 2011
    Co-Authors: Shigeru Kitani, Satoshi Omura, Miho Uchida, Satoshi Takamatsu, Kiyoko T Miyamoto, Tohru Nagamitsu, Elisa Herawati, Hiroyuki Iguchi, Kouhei Nishitomi, Haruo Ikeda
    Abstract:

    Gram-positive bacteria of the genus Streptomyces are industrially important microorganisms, producing >70% of commercially important antibiotics. The production of these compounds is often regulated by low-molecular-weight bacterial hormones called autoregulators. Although 60% of Streptomyces strains may use γ-butyrolactone–type molecules as autoregulators and some use furan-type molecules, little is known about the signaling molecules used to regulate antibiotic production in many other members of this genus. Here, we purified a signaling molecule (avenolide) from Streptomyces avermitilis—the producer of the important anthelmintic agent avermectin with annual world sales of $850 million—and determined its structure, including stereochemistry, by spectroscopic analysis and chemical synthesis as (4S,10R)-10-hydroxy-10-methyl-9-oxo-dodec-2-en-1,4-olide, a class of Streptomyces autoregulator. Avenolide is essential for eliciting avermectin production and is effective at nanomolar concentrations with a minimum effective concentration of 4 nM. The aco gene of S. avermitilis, which encodes an acyl-CoA oxidase, is required for avenolide biosynthesis, and homologs are also present in Streptomyces fradiae, Streptomyces ghanaensis, and Streptomyces griseoauranticus, suggesting that butenolide-type autoregulators may represent a widespread and another class of Streptomyces autoregulator involved in regulating antibiotic production.

  • Geosmin Biosynthesis in Streptomyces avermitilis . Molecular Cloning, Expression, and Mechanistic Study of the Germacradienol/Geosmin Synthase
    The Journal of Antibiotics, 2006
    Co-Authors: David E Cane, Seiji Kobayashi, Xiaofei He, Satoshi Omura, Haruo Ikeda
    Abstract:

    Geosmin Biosynthesis in Streptomyces avermitilis . Molecular Cloning, Expression, and Mechanistic Study of the Germacradienol/Geosmin Synthase

  • Pentalenolactone biosynthesis. Molecular cloning and assignment of biochemical function to PtlH, a non-heme iron dioxygenase of Streptomyces avermitilis.
    Journal of the American Chemical Society, 2006
    Co-Authors: Zheng You, Satoshi Omura, Haruo Ikeda, David E Cane
    Abstract:

    The hydroxylase encoded by the ptlH (SAV2991) gene from the pentalenolactone gene cluster of Streptomyces avermitilis was cloned by PCR and expressed in Escherichia coli as an N-terminal His6-tag protein. Incubation of recombinant PtlH with (±)-1-deoxypentalenic acid (5) in the presence of Fe(II), α-ketoglutarate, and O2 gave (−)-11β-hydroxy-1-deoxypentalenic acid (8), whose structure and stereochemistry were determined by a combination of 1H, 13C, COSY, HMQC, HMBC, and NOESY NMR. The steady-state kinetic parameters were kcat = 4.2 ± 0.6 s-1 and Km (5) = 0.57 ± 0.19 mM. 8 is a new intermediate in the conversion of the sesquiterpene pentalenene (3) to pentalenolactone (1).

Jidong Wang - One of the best experts on this subject based on the ideXlab platform.

Byung-gee Kim - One of the best experts on this subject based on the ideXlab platform.

  • Regioselective Biotransformation of Phloretin Using Streptomyces avermitilis MA4680
    Biotechnology and Bioprocess Engineering, 2020
    Co-Authors: Woo-il Kim, Jong-ki Lee, Kwon-young Choi, Byung-gee Kim, June-hyung Kim
    Abstract:

    Streptomyces avermitilis MA4680 was used for the biotransformation of phloretin, one of the dihydrochalcone found in apple bark. After LC analysis, two main biotransformed products were identified and they were further analyzed using GC/MS. After BSTFA derivatization of biotransformed product, they were interpreted as regioselectively hydroxylated products of phloretin in Bring. Maximum conversion of phloretin was 6.7% with 1 h of reaction, and the phloretin and reaction products were completely metabolized after 3 h of reaction due to cellular metabolism. Addition of 0.5 mM quinidine completely blocked hydroxylation of phloretin, which means the hydroxylation proceed by P450 monoxygenase dependent metabolism. Addition of Brij 35 detergent resulted in 150% increase of hydroxylated product due to facilitating transport of phloretin and its reaction product across cellular membrane.

  • Regioselective hydroxylation of trans-resveratrol via inhibition of tyrosinase from Streptomyces avermitilis MA4680.
    ACS chemical biology, 2012
    Co-Authors: Nahum Lee, Eun Jung Kim, Byung-gee Kim
    Abstract:

    Secreted tyrosinase from melanin-forming Streptomyces avermitilis MA4680 was involved in both ortho-hydroxylation and further oxidation of trans-resveratrol, leading to the formation of melanin. This finding was confirmed by constructing deletion mutants of melC2 and melD2 encoding extracellular and intracellular tyrosinase, respectively; the melC2 deletion mutant did not produce piceatannol as well as melanin, whereas the melD2 deletion mutant oxidized resveratrol and synthesized melanin with the same yields, suggesting that MelC2 is responsible for ortho-hydroxylation of resveratrol. Extracellular tyrosinase (MelC2) efficiently converted trans-resveratrol into piceatannol in the presence of either tyrosinase inhibitors or reducing agents such as catechol, NADH, and ascorbic acid. Reducing agents slow down the dioxygenase reaction of tyrosinase. In the presence of catechol, the regio-specific hydroxylation of trans-resveratrol was successfully performed by whole cell biotransformation, and further oxidat...

  • regioselective hydroxylation of daidzein using p450 cyp105d7 from Streptomyces avermitilis ma4680
    Biotechnology and Bioengineering, 2010
    Co-Authors: Bishnu Prasad Pandey, Kwon-young Choi, Changhyun Roh, Nahum Lee, Eun Jung Kim, Taejin Kim, Hyundon Yun, Byung-gee Kim
    Abstract:

    Regiospecific 3'-hydroxylation reaction of daidzein was performed with CYP105D7 from Streptomyces avermitilis MA4680 expressed in Escherichia coli. The apparent K(m) and k(cat) values of CYP105D7 for daidzein were 21.83 +/- 6.3 microM and 15.01 +/- 0.6 min(-1) in the presence of 1 microM of CYP105D7, putidaredoxin (CamB) and putidaredoxin reductase (CamA), respectively. When CYP105D7 was expressed in S. avermitilis MA4680, its cytochrome P450 activity was confirmed by the CO-difference spectra at 450 nm using the whole cell extract. When the whole-cell reaction for the 3'-hydroxylation reaction of daidzein was carried out with 100 microM of daidzein in 100 mM of phosphate buffer (pH 7.5), the recombinant S. avermitilis grown in R2YE media overexpressing CYP105D7 and ferredoxin FdxH (SAV7470) showed a 3.6-fold higher conversion yield (24%) than the corresponding wild type cell (6.7%). In a 7 L (working volume 3 L) jar fermentor, the recombinants S. avermitilis grown in R2YE media produced 112.5 mg of 7,3',4'-trihydroxyisoflavone (i.e., 29.5% conversion yield) from 381 mg of daidzein in 15 h.

  • Regioselective hydroxylation of isoflavones by Streptomyces avermitilis MA-4680.
    Journal of bioscience and bioengineering, 2009
    Co-Authors: Changhyun Roh, Kwon-young Choi, June-hyung Kim, Su-hyun Seo, Min-ho Cha, Bishnu Prasad Pandey, Jun-seong Park, Duck Hee Kim, Ih Seop Chang, Byung-gee Kim
    Abstract:

    Abstract Screening of bacterial whole cells was performed for regioselective hydroxylation of daidzein and genistein. Among the strains examined, Streptomyces avermitilis MA-4680 showed high ortho-dihydroxylation activity to produce 3′,4′,7-trihydroxyisoflavone and 3′,4′,5,7-tetrahydroxyisoflavone from daidzein (4′,7-dihydroxyisoflavone) and genistein (4′,5,7-trihydroxyisoflavone), respectively. Using 100 mg cells (wet wt.) and 1% (v/v) Triton X100 in 1 ml of total reaction volume, where 100 μl of the substrate solution (0.5 mM in 10% (v/v) mixed solvent of DMSO:MeOH = 3:7) was added to 900 μl of potassium phosphate buffer (100 mM, pH 7.2), a 16% molar conversion yield of 3′,4′,7-trihydroxyisoflavone was obtained from 0.5 mM daidzein after 24 h of reaction time at 28 °C and 200 rpm. Ketoconazole significantly (ca. 90%) inhibited the ortho-hydroxylation activity of daidzein, suggesting that cytochrome P450 enzymes putatively play roles in regiospecific daidzein hydroxylation. The analysis of the reaction products was determined by gas chromatography/mass spectrometry (GC/MS) and 1 H NMR.

David E Cane - One of the best experts on this subject based on the ideXlab platform.