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ABSTRACT: Cytochrome P450foxy (P450foxy) is a fatty acid (FA) monooxygenase that is characterized by self-sufficient catalysis and high turnover numbers due to the fused structure of cytochrome P450 and its reductase. Here we found that resting recombinant Escherichia coli cells producing P450foxy converted saturated FA with a chain length of 7-16 carbon atoms to their omega-1 to omega-3 hydroxy derivatives. Most products were recovered from the culture supernatant. Decanoic acid was most efficiently converted to omega-1 to omega-3 hydroxy decanoic acids in the order of omega-1>omega-2>omega-3, with a total product yield of 47%. We also found that P450foxy was more active against physiological fatty acyl esters such as monopalmitoyl glycerol, monopalmitoyl phospholipid, and palmitoyl CoA than free palmitic acid. The bacteria producing P450foxy were applicable as biocatalysts in the production of omega-1 hydroxy palmitic acid from lard, vegetable, and soy sauce oil wastes from the food industry.
Applied Microbiology and Biotechnology 08/2008; 79(6):981-8. · 3.42 Impact Factor
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ABSTRACT: The major sex pheromone components of Drosophila ananassae and D. pallidosa, (Z,Z)-5,25-hentriacontadiene and (Z,Z)-5,27-tritriacontadiene, respectively, were synthesized by using the Wittig olefination and sulfone coupling reactions as the C-C bond-forming steps.
Bioscience Biotechnology and Biochemistry 09/2005; 69(8):1620-3. · 1.28 Impact Factor
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ABSTRACT: 4,8-Dimethyldecanal (4,8-DMD) is the aggregation pheromone produced by male red flour beetles (RFB), Tribolium castaneum. To elucidate the biosynthetic origin of 4,8-DMD, the following studies were performed: (1) effects of juvenile hormone (JH) III, and pathway inhibitors mevastatin, an inhibitor of the mevalonate pathway, and 2-octynoic acid, an inhibitor of the fatty acid pathway, were tested to determine whether 4,8-DMD is derived from the fatty acid pathway or the mevalonate pathway; (2) incorporation of 13C-labeled acetate, propionate, and mevalonolactone into 4,8-DMD was measured to directly determine the biosynthetic origin of 4,8-DMD; and (3) incorporation of deuterium-labeled precursors, including 2-methylbutanoate (C5D), 4-methylhexanoate (C7D), 2,6-dimethyloctanoate (C10D), and 4,8-dimethyldecanoate (C12D) was tested to determine whether 4,8-DMD is biosynthesized in the sequence Ac-Pr-Ac-Pr-Ac (Ac; acetate, Pr; propionate). JH III was topically applied to males at various doses. Inhibitors and isotopically labeled substrates were administered orally by feeding the beetles flour treated with the substrates of interest, after which volatiles were collected from both sexes of RFBs. The amount of 4,8-DMD produced was significantly reduced with increasing doses of JH III. Also, 2-octynoic acid inhibited the production of 4,8-DMD, but mevastatin did not. Exposure of RFBs to [1-(13)C]acetate and [1-(13)C]propionate, but not [2-(13)C]mevalonolactone, resulted in incorporation of the labeled compounds into 4,8-DMD. RFBs fed flour treated with deuterium-labeled C5D, C10D, and C12D, but not C7D, incorporated these compounds into 4,8-DMD. The findings that the production of 4,8-DMD was inhibited by 2-octynoic acid but unaffected by mevastatin, combined with the high incorporation of [1-(13)C]acetate and [1-(13)C]propionate into 4,8-DMD and the incorporation of deuterated precursors, unambiguously demonstrated that 4,8-DMD is of fatty acid rather than terpene biosynthetic origin, and that the biosynthesis of 4,8-DMD proceeds in the sequence Ac-Pr-Ac-Pr-Ac.
Journal of Chemical Ecology 07/2005; 31(6):1381-400. · 2.66 Impact Factor
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ABSTRACT: The anal secretions of 16 phlaeothripine thrips species (Thysanoptera: Phlaeothripidae) were studied, including a reinvestigation of three species previously reported. A total of 37 components were detected, including hydrocarbons, acetates, terpenes, carboxylic acids, a quinone, an aromatic compound, and a pyranone compound. The secretions of all species were composed of some of these components, with Xylaplothrips inquilinus possessing as many as 11 components. Of these components, (Z)-9-octadecene, (Z)-9-nonadecene, nonadecadiene, octanoic acid, decanoic acid, geranial, neral, alpha-pinene, beta-pinene, caryophyllene, 2-hydroxy-6-methylbenzaldehyde, and two unidentified monoterpenes [UK-I (M+136) and UK-II (M+168)] were detected for the first time. The chemicals were species-specific; four Liothrips species and three Holothrips species could be distinguished from each other and their congeners by the GC profiles of the ether extracts of their anal secretions. The anal secretions of gall-inducing thrips commonly contained terpenes. of which citral (a mixture of geranial and neral) and beta-acaridial repelled ants or had antifungal activity. The findings suggest that these terpenes play a defensive role and prevent galls from fungal infestation. 3-Butanoyl-4-hydroxy-6-methyl-2H-pyran-2-one, found from three Holothrips spp., caused paralysis in ants. Chemical analysis of anal secretion components is a useful method for the classification of tubuliferan species that are difficult to distinguish on the basis of morphological characters.
Journal of Chemical Ecology 03/2004; 30(2):409-23. · 2.66 Impact Factor
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ABSTRACT: Host-recognizing kairomones for the stinging behavior of the parasitic wasp, Anisopteromalus calandrae, were identified on host azuki bean weevil larvae, Callosobruchus chinensis (L.). The kairomones were extracted with acetone from Chinese green beans, from which emerged wasps and host weevils had been removed. The kairomones are a mixture of triacylglycerols and fatty acids, each of which is separately active, and with no observable synergistic effect between them. These compounds are known to be constituents of an oviposition-marking pheromone of host azuki bean weevils. However, they differ from the previously reported saturated hydrocarbons and diacylglycerols of the kairomone that another parasitic wasp, Dinarmus basalis, uses for the host recognition of C. chinensis. Thus, A. calandrae and D. basalis selectively utilize different constituents of the oviposition-marking pheromone of C. chinensis as host-recognizing kairomones.
Journal of Chemical Ecology 07/2002; 28(6):1209-20. · 2.66 Impact Factor
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ABSTRACT: Cytochrome P450foxy (P450foxy, CYP505) is a fused protein of cytochrome P450 (P450) and its reductase isolated from the fungus Fusarium oxysporum, which catalyzes the subterminal (omega-1 approximately omega-3) hydroxylation of fatty acids. Here, we produced, purified and characterized a fused recombinant protein (rP450foxy) using the Escherichia coli expression system. Purified rP450foxy was catalytically and spectrally indistinguishable from the native protein, but most of the rP450foxy was recovered in the soluble fraction of E. coli cells unlike the membrane-bound native protein. The results are consistent with our notion that the native protein is targeted to the membrane by a post-translational modification mechanism. We also discovered that P450foxy could use shorter saturated fatty acid chains (C9 and C10) as a substrate. The regiospecificity (omega-1 approximately omega-3) of hydroxylation due to the enzymatic reaction for the short substrates (decanoate, C10; undecanoate, C11) was the same as that for longer substrates. Steady state kinetic studies showed that the kcat values for all substrates tested (C9-C16) were of the same magnitude (1200-1800 min-1), whereas the catalytic efficiency (kcat/Km) was higher for longer fatty acids. Substrate inhibition was observed with fatty acid substrates longer than C13, and the degree of inhibition increased with increasing chain length. This substrate inhibition was not apparent with P450BM3, a bacterial counterpart of P450foxy, which was the first obvious difference in their catalytic properties to be identified. Kinetic data were consistent with the inhibition due to binding of the second substrate. We discuss the inhibition mechanism based on differences between P450foxy and P450BM3 in key amino acid residues for substrate binding.
European Journal of Biochemistry 05/2002; 269(8):2075-82. · 3.58 Impact Factor
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ABSTRACT: A host-recognizing kairomone responsible for the stinging behavior of the parasitic wasp, Dinarmus basalis, was studied. Fresh azuki beans coated with an acetone extract of the azuki beans, from which both emerged wasps and their host weevils were removed, elicited stinging behavior from female wasps. The kairomone is a mixture of saturated hydrocarbons and diacylglycerols, both of which are required for activity. The kairomone is composed of normal and methyl-branched hydrocarbons with carbon numbers ranging from 25 to 35, most of which are known as the hydrocarbon constituents of an oviposition-marking pheromone of the host azuki bean weevils, Callosobruchus chinensis. This indicates that D. basalis utilizes the oviposition-marking pheromone of its host weevils as a host-recognizing kairomone.
Journal of Chemical Ecology 11/2000; 26(12):2677-2695. · 2.66 Impact Factor
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ABSTRACT: Anal secretions of 17 species in nine genera of the subfamily Idolothripinae were analzyed by TLC, GC, and GC-MS. From all species, juglone and two or more carboxylic acids with carbon numbers ranging from 10 to 14 were detected; the exception was Gastrothrips sp., from which none was detected. Twelve carboxylic acids were found, of which decanoic acid, (E)-4-decenoic acid, and (Z)-5-tetradecenoic acid were widely found, and acid ingredients were fairly species-specific. The Gastrothrips sp. was the only exception, and it had no secretion components among the tubuliferan species so far analyzed. Juglone is thought to be a potent defensive allomone, and roles of the carboxylic acids are discussed.
Journal of Chemical Ecology 09/2000; 26(10):2449-2458. · 2.66 Impact Factor
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ABSTRACT: Fatty acid (ω−1–ω−3) hydroxylase of Fusarium oxysporum (P450foxy) was produced in yeast. The recombinant protein showed a close resemblance, in the spectral and in the enzymatic properties, to native P450foxy. P450foxy is also similar in their self-sufficiency and in the monooxygenase activity to P450BM3 from Bacillus megaterium, a bacterial counterpart of P450foxy. However, significant differences in their substrate specificity, subcellular localization, and inducing patterns are found between these two cytochromes P450.
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