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Dive into the research topics where Futoshi Taura is active.

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Featured researches published by Futoshi Taura.


FEBS Letters | 2007

Cannabidiolic-acid synthase, the chemotype-determining enzyme in the fiber-type Cannabis sativa.

Futoshi Taura; Supaart Sirikantaramas; Yoshinari Shoyama; Kazuyoshi Yoshikai; Yukihiro Shoyama; Satoshi Morimoto

Cannabidiolic‐acid (CBDA) synthase is the enzyme that catalyzes oxidative cyclization of cannabigerolic‐acid into CBDA, the dominant cannabinoid constituent of the fiber‐type Cannabis sativa. We cloned a novel cDNA encoding CBDA synthase by reverse transcription and polymerase chain reactions with degenerate and gene‐specific primers. Biochemical characterization of the recombinant enzyme demonstrated that CBDA synthase is a covalently flavinylated oxidase. The structural and functional properties of CBDA synthase are quite similar to those of tetrahydrocannabinolic‐acid (THCA) synthase, which is responsible for the biosynthesis of THCA, the major cannabinoid in drug‐type Cannabis plants.


Journal of Biological Chemistry | 1996

Purification and Characterization of Cannabidiolic-acid Synthase from Cannabis sativa L. BIOCHEMICAL ANALYSIS OF A NOVEL ENZYME THAT CATALYZES THE OXIDOCYCLIZATION OF CANNABIGEROLIC ACID TO CANNABIDIOLIC ACID

Futoshi Taura; Satoshi Morimoto; Yukihiro Shoyama

We identified a unique enzyme that catalyzes the oxidocyclization of cannabigerolic acid to cannabidiolic acid (CBDA) in Cannabis sativa L. (CBDA strain). The enzyme, named CBDA synthase, was purified to apparent homogeneity by a four-step procedure: ammonium sulfate precipitation followed by chromatography on DEAE-cellulose, phenyl-Sepharose CL-4B, and hydroxylapatite. The active enzyme consists of a single polypeptide with a molecular mass of 74 kDa and a pI of 6.1. The NH2-terminal amino acid sequence of CBDA synthase is similar to that of Δ1-tetrahydrocannabinolic-acid synthase. CBDA synthase does not require coenzymes, molecular oxygen, hydrogen peroxide, and metal ion cofactors for the oxidocyclization reaction. These results indicate that CBDA synthase is neither an oxygenase nor a peroxidase and that the enzymatic cyclization does not proceed via oxygenated intermediates. CBDA synthase catalyzes the formation of CBDA from cannabinerolic acid as well as cannabigerolic acid, although the kcat for the former (0.03 s−1) is lower than that for the latter (0.19 s−1). Therefore, we conclude that CBDA is predominantly biosynthesized from cannabigerolic acid rather than cannabinerolic acid.


Journal of Biological Chemistry | 1998

Novel hydrogen peroxide metabolism in suspension cells of Scutellaria baicalensis Georgi

Satoshi Morimoto; Norifumi Tateishi; Tomoko Matsuda; Hiroyuki Tanaka; Futoshi Taura; Naruto Furuya; Nobuaki Matsuyama; Yukihiro Shoyama

We identified a rapid and novel system to effectively metabolize a large amount of H2O2 in the suspension cells ofScutellaria baicalensis Georgi. In response to an elicitor, the cells immediately initiate the hydrolysis of baicalein 7-O-β-d-glucuronide by β-glucuronidase, and the released baicalein is then quickly oxidized to 6,7-dehydrobaicalein by peroxidases. Hydrogen peroxide is effectively consumed during the peroxidase reaction. The β-glucuronidase inhibitor, saccharic acid 1,4-lactone, significantly reduced the H2O2-metabolizing ability of theScutellaria cells, indicating that β-glucuronidase, which does not catalyze the H2O2 degradation, plays an important role in the H2O2 metabolism. As H2O2-metabolizing enzymes, we purified two peroxidases using ammonium sulfate precipitation followed by sequential chromatography on CM-cellulose and hydroxylapatite. Both peroxidases show high H2O2-metabolizing activity using baicalein, whereas other endogenous flavones are not substrates of the peroxidase reaction. Therefore, baicalein predominantly contributed to H2O2 metabolism. Because β-glucuronidase, cell wall peroxidases, and baicalein pre-exist inScutellaria cells, their constitutive presence enables the cells to rapidly induce the H2O2-metabolizing system.


FEBS Letters | 2009

Characterization of olivetol synthase, a polyketide synthase putatively involved in cannabinoid biosynthetic pathway.

Futoshi Taura; Shinji Tanaka; Chiho Taguchi; Tomohide Fukamizu; Hiroyuki Tanaka; Yukihiro Shoyama; Satoshi Morimoto

Alkylresorcinol moieties of cannabinoids are derived from olivetolic acid (OLA), a polyketide metabolite. However, the polyketide synthase (PKS) responsible for OLA biosynthesis has not been identified. In the present study, a cDNA encoding a novel PKS, olivetol synthase (OLS), was cloned from Cannabis sativa. Recombinant OLS did not produce OLA, but synthesized olivetol, the decarboxylated form of OLA, as the major reaction product. Interestingly, it was also confirmed that the crude enzyme extracts from flowers and rapidly expanding leaves, the cannabinoid‐producing tissues of C. sativa, also exhibited olivetol‐producing activity, suggesting that the native OLS is functionally expressed in these tissues. The possibility that OLS could be involved in OLA biosynthesis was discussed based on its catalytic properties and expression profile.


Phytochemistry | 1998

Purification and characterization of cannabichromenic acid synthase from Cannabis sativa

Satoshi Morimoto; Kengo Komatsu; Futoshi Taura; Yukihiro Shoyama

Cannabichromenic acid synthase was purified to apparent homogeneity by sequential column chromatography including DEAE-cellulose, phenyl-Sepharose CL-4B, and hydroxylapatite. The enzyme catalysed the oxidocyclization of cannabigerolic acid and cannabinerolic acid to cannabichromenic acid. The K(m) values for both substrates were in the same order of magnitude although the Vmax value for the former was higher than that for the latter. These results suggested that cannabichromenic acid is predominantly formed from cannabigerolic acid rather than cannabinerolic acid. The enzyme required neither molecular oxygen nor hydrogen peroxide, indicating that the cannabichromenic acid synthase reaction proceeds through direct dehydrogenation without hydroxylation.


BMC Plant Biology | 2004

Molecular cloning and functional expression of geranylgeranyl pyrophosphate synthase from Coleus forskohlii Briq

Surang Engprasert; Futoshi Taura; Makoto Kawamukai; Yukihiro Shoyama

BackgroundIsopentenyl diphosphate (IPP), a common biosynthetic precursor to the labdane diterpene forskolin, has been biosynthesised via a non-mevalonate pathway. Geranylgeranyl diphosphate (GGPP) synthase is an important branch point enzyme in terpenoid biosynthesis. Therefore, GGPP synthase is thought to be a key enzyme in biosynthesis of forskolin. Herein we report the first confirmation of the GGPP synthase gene in Coleus forskohlii Briq.ResultsThe open reading frame for full-length GGPP synthase encodes a protein of 359 amino acids, in which 1,077 nucleotides long with calculated molecular mass of 39.3 kDa. Alignments of C. forskohlii GGPP synthase amino acid sequences revealed high homologies with other plant GGPP synthases. Several highly conserved regions, including two aspartate-rich motifs were identified. Transient expression of the N-terminal region of C. forskohlii GGPP synthase-GFP fusion protein in tobacco cells demonstrated subcellular localization in the chloroplast. Carotenoid production was observed in Escherichia coli harboring pACCAR25ΔcrtE from Erwinia uredovora and plasmid carrying C. forskohlii GGPP synthase. These results suggested that cDNA encoded functional GGPP synthase. Furthermore, C. forskohlii GGPP synthase expression was strong in leaves, decreased in stems and very little expression was observed in roots.ConclusionThis investigation proposed that forskolin was synthesised via a non-mevalonate pathway. GGPP synthase is thought to be involved in the biosynthesis of forskolin, which is primarily synthesised in the leaves and subsequently accumulates in the stems and roots.


Current Pharmaceutical Biotechnology | 2007

Recent Advances in Cannabis sativa Research: Biosynthetic Studies and Its Potential in Biotechnology

Supaart Sirikantaramas; Futoshi Taura; Satoshi Morimoto; Yukihiro Shoyama

Cannabinoids, consisting of alkylresorcinol and monoterpene groups, are the unique secondary metabolites that are found only in Cannabis sativa. Tetrahydrocannabinol (THC), cannabidiol (CBD) and cannabichromene (CBC) are well known cannabinoids and their pharmacological properties have been extensively studied. Recently, biosynthetic pathways of these cannabinoids have been successfully established. Several biosynthetic enzymes including geranylpyrophosphate:olivetolate geranyltransferase, tetrahydrocannabinolic acid (THCA) synthase, cannabidiolic acid (CBDA) synthase and cannabichromenic acid (CBCA) synthase have been purified from young rapidly expanding leaves of C. sativa. In addition, molecular cloning, characterization and localization of THCA synthase have been recently reported. THCA and cannabigerolic acid (CBGA), its substrate, were shown to be apoptosis-inducing agents that might play a role in plant defense. Transgenic tobacco hairy roots expressing THCA synthase can produce THCA upon feeding of CBGA. These results open the way for biotechnological production of cannabinoids in the future.


Journal of Biological Chemistry | 2007

Identification and Characterization of Cannabinoids That Induce Cell Death through Mitochondrial Permeability Transition in Cannabis Leaf Cells

Satoshi Morimoto; Yumi Tanaka; Kaori Sasaki; Hiroyuki Tanaka; Tomohide Fukamizu; Yoshinari Shoyama; Yukihiro Shoyama; Futoshi Taura

Cannabinoids are secondary metabolites stored in capitate-sessile glands on leaves of Cannabis sativa. We discovered that cell death is induced in the leaf tissues exposed to cannabinoid resin secreted from the glands, and identified cannabichromenic acid (CBCA) and Δ1-tetrahydrocannabinolic acid (THCA) as unique cell death mediators from the resin. These cannabinoids effectively induced cell death in the leaf cells or suspension-cultured cells of C. sativa, whereas pretreatment with the mitochondrial permeability transition (MPT) inhibitor cyclosporin A suppressed this cell death response. Examinations using isolated mitochondria demonstrated that CBCA and THCA mediate opening of MPT pores without requiring Ca2+ and other cytosolic factors, resulting in high amplitude mitochondrial swelling, release of mitochondrial proteins (cytochrome c and nuclease), and irreversible loss of mitochondrial membrane potential. Therefore, CBCA and THCA are considered to cause serious damage to mitochondria through MPT. The mitochondrial damage was also confirmed by a marked decrease of ATP level in cannabinoid-treated suspension cells. These features are in good accord with those of necrotic cell death, whereas DNA degradation was also observed in cannabinoid-mediated cell death. However, the DNA degradation was catalyzed by nuclease(s) released from mitochondria during MPT, indicating that this reaction was not induced via a caspase-dependent apoptotic pathway. Furthermore, the inhibition of the DNA degradation only slightly blocked the cell death induced by cannabinoids. Based on these results, we conclude that CBCA and THCA have the ability to induce necrotic cell death via mitochondrial dysfunction in the leaf cells of C. sativa.


Journal of Biological Chemistry | 1999

Identification and Molecular Characterization of Novel Peroxidase with Structural Protein-like Properties

Satoshi Morimoto; Norifumi Tateishi; Masako Inuyama; Futoshi Taura; Hiroyuki Tanaka; Yukihiro Shoyama

Elicitor treatment or mechanical damage toScutellaria baicalensis Georgi (skullcap plants) callus causes an immediate insolubilization of a 36-kDa protein into cell walls. The 36-kDa protein was identified as peroxidase 1 by analysis of its internal amino acid sequence and by immunoblotting using affinity-purified anti-peroxidase 1. Insolubilized peroxidase 1 is cross-linked to lignin through covalent bonds, and the cross-linking is catalyzed in the presence of H2O2 by peroxidase 1 itself. The properties of insolubilized peroxidase 1 resemble those of defense-related structural proteins (extensins and proline-rich proteins) cross-linked to cell wall. Although the isozymes peroxidases 2 and 3 have enzyme activities similar to peroxidase 1, they are not insolubilized by stress treatment. Molecular characterization established that peroxidase 1 contains regions characteristic of structural proteins, but peroxidases 2 and 3 do not have such regions. These results suggest that among the three isozymes, only peroxidase 1 has a structural protein-like function as well as an enzymatic function.


Acta Crystallographica Section F-structural Biology and Crystallization Communications | 2005

Crystallization of Δ1-tetrahydrocannabinolic acid (THCA) synthase from Cannabis sativa

Yoshinari Shoyama; Ayako Takeuchi; Futoshi Taura; Taro Tamada; Motoyasu Adachi; Ryota Kuroki; Yukihiro Shoyama; Satoshi Morimoto

Delta1-Tetrahydrocannabinolic acid (THCA) synthase is a novel oxidoreductase that catalyzes the biosynthesis of the psychoactive compound THCA in Cannabis sativa (Mexican strain). In order to investigate the structure-function relationship of THCA synthase, this enzyme was overproduced in insect cells, purified and finally crystallized in 0.1 M HEPES buffer pH 7.5 containing 1.4 M sodium citrate. A single crystal suitable for X-ray diffraction measurement was obtained in 0.09 M HEPES buffer pH 7.5 containing 1.26 M sodium citrate. The crystal diffracted to 2.7 A resolution at beamline BL41XU, SPring-8. The crystal belonged to the primitive cubic space group P432, with unit-cell parameters a = b = c = 178.2 A. The calculated Matthews coefficient was approximately 4.1 or 2.0 A3 Da(-1) assuming the presence of one or two molecules of THCA synthase in the asymmetric unit, respectively.

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Yukihiro Shoyama

Nagasaki International University

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Ryota Kuroki

Japan Atomic Energy Research Institute

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