Hor-Gil Hur

Enantioselective synthesis of S-equol from dihydrodaidzein by a newly isolated anaerobic human intestinal bacterium

ABSTRACT A newly isolated rod-shaped, gram-negative anaerobic bacterium from human feces, named Julong 732, was found to be capable of metabolizing the isoflavone dihydrodaidzein to S-equol under anaerobic conditions. The metabolite, equol, was identified by using electron impact ionization mass spectrometry, 1H and 13C nuclear magnetic resonance spectroscopy, and UV spectral analyses. However, strain Julong 732 was not able to produce equol from daidzein, and tetrahydrodaidzein and dehydroequol, which are most likely intermediates in the anaerobic metabolism of dihydrodaidzein, were not detected in bacterial culture medium containing dihydrodaidzein. Chiral stationary-phase high-performance liquid chromatography eluted only one metabolite, S-equol, which was produced from a bacterial culture containing a racemic mixture of dihydrodaidzein. Strain Julong 732 did not show racemase activity to transform R-equol to S-equol and vice versa. Its full 16S rRNA gene sequence (1,429 bp) had 92.8% similarity to that of Eggerthella hongkongenis HKU10. This is the first report of a single bacterium capable of converting a racemic mixture of dihydrodaidzein to enantiomeric pure S-equol.

Growth Mechanism of Amorphous Selenium Nanoparticles Synthesized by Shewanella sp. HN-41

Shewanella sp. HN-41 was exploited for selenium nanoparticles synthesis from aqueous selenite compounds under anaerobic conditions. Various reaction conditions, including reaction time, initial biomass, and initial selenite concentration, were systematically investigated to determine their effects on particle size distribution and formation rate. The biomass concentration of Shewanella sp. HN-41 had no significant effect on average particle size but strongly influenced reduction rate and size distribution. Initial selenite concentration (0.01–1.0 mM) also had no significant effect on the average particle size, but affected the early growth stage mechanism of selenium particle production, which was modeled using a Michaelis Menten model. The HR-TEM and SAED patterns indicated that the synthesized selenium nanoparticles were amorphous.

Absence of Escherichia coli Phylogenetic Group B2 Strains in Humans and Domesticated Animals from Jeonnam Province, Republic of Korea

ABSTRACT Multiplex PCR analyses of DNAs from genotypically unique Escherichia coli strains isolated from the feces of 138 humans and 376 domesticated animals from Jeonnam Province, South Korea, performed using primers specific for the chuA and yjaA genes and an unknown DNA fragment, TSPE4.C2, indicated that none of the strains belonged to E. coli phylogenetic group B2. In contrast, phylogenetic group B2 strains were detected in about 17% (8 of 48) of isolates from feces of 24 wild geese and in 3% (3 of 96) of isolates obtained from the Yeongsan River in Jeonnam Province, South Korea. The distribution of E. coli strains in phylogenetic groups A, B1, and D varied depending on the host examined, and there was no apparent seasonal variation in the distribution of strains in phylogenetic groups among the Yeongsan River isolates. The distribution of four virulence genes (eaeA, hlyA, stx1, and stx2) in isolates was also examined by using multiplex PCR. Virulence genes were detected in about 5% (38 of 707) of the total group of unique strains examined, with 24, 13, 13, and 9 strains containing hlyA, eaeA, stx2, and stx1, respectively. The virulence genes were most frequently present in phylogenetic group B1 strains isolated from beef cattle. Taken together, results of these studies indicate that E. coli strains in phylogenetic group B2 were rarely found in humans and domesticated animals in Jeonnam Province, South Korea, and that the majority of strains containing virulence genes belonged to phylogenetic group B1 and were isolated from beef cattle. Results of this study also suggest that the relationship between the presence and types of virulence genes and phylogenetic groupings may differ among geographically distinct E. coli populations.

Organic Acid-Dependent Iron Mineral Formation by a Newly Isolated Iron-Reducing Bacterium, Shewanella sp. HN-41

The influence of organic acids used as substrates for bacterial respiration coupled to dissimilatory Fe(III) reduction on the mineralogy of formed iron minerals was examined. A newly isolated dissimilatory iron-reducing bacterium, Shewanella strain HN-41 exhibited different mineral formation patterns resulting from the reduction of poorly crystalline Fe(III)-oxyhydroxide, akaganeite (β-FeOOH; ∼ 70 mM) depending upon the presence of the different organic acids: lactate, pyruvate, and formate (10 mM) under anaerobic conditions. X-ray diffraction analysis identified the minerals as magnetite, siderite, and a mixture of magnetite and siderite, which were produced by strain HN-41 using lactate, pyruvate, and formate as a sole electron donor, respectively. With the descending order of the amount of Fe(II) in aqueous phase, the pyruvate-, formate-, and lactate-incubations released Fe(II) into aqueous phase from the poorly crystalline akaganeite by the bacterial Fe(III) reduction. The amount of Fe(II) released into aqueous phase was inversely related to the degree of crystallinity. Magnetite produced during lactate-incubation with the least amount of Fe(II) in aqueous phase, and final pH of 8.6 and Eh of –408 mV showed the highest crystallinity, while siderite formed by pyruvate-incubation with the largest release of Fe(II) in aqueous phase, and final pH of 8.1 and Eh of –394 mV showed the lowest crystallinity. When strain HN-41 was incubated with the organic acid substrates at 10 mM and Fe(III)-citrate at 20 mM, total inorganic carbon (TIC) contents, which is the collective term of free carbon dioxide [CO 2 ], carbonate ion [CO3 2 − ], bicarbonate ion [HCO 3 −], and carbonic acid [H 2 CO 3 ], also showed different trends with different organic acids as electron donors. TICs produced from both pyruvate-and formate-incubations were relatively fast and decreased with time after early phase increases, while TIC from the lactate-incubation increased gradually. Therefore, low molecular weight organic acids lactate, pyruvate, and formate, may differentially affect mineral formation through chemistry change of the environmental conditions created by the newly isolated dissimilatory iron-reducing bacterium Shewanella strain HN-41.

Molecular Characterization of Bacteriophages for Microbial Source Tracking in Korea

ABSTRACT We investigated coliphages from various fecal sources, including humans and animals, for microbial source tracking in South Korea. Both somatic and F+-specific coliphages were isolated from 43 fecal samples from farms, wild animal habitats, and human wastewater plants. Somatic coliphages were more prevalent and abundant than F+ coliphages in all of the tested fecal samples. We further characterized 311 F+ coliphage isolates using RNase sensitivity assays, PCR and reverse transcription-PCR, and nucleic acid sequencing. Phylogenetic analyses were performed based on the partial nucleic acid sequences of 311 F+ coliphages from various sources. F+ RNA coliphages were most prevalent among geese (95%) and were least prevalent in cows (5%). Among the genogroups of F+ RNA coliphages, most F+ coliphages isolated from animal fecal sources belonged to either group I or group IV, and most from human wastewater sources were in group II or III. Some of the group I coliphages were present in both human and animal source samples. F+ RNA coliphages isolated from various sources were divided into two main clusters. All F+ RNA coliphages isolated from human wastewater were grouped with Qβ-like phages, while phages isolated from most animal sources were grouped with MS2-like phages. UniFrac significance statistical analyses revealed significant differences between human and animal bacteriophages. In the principal coordinate analysis (PCoA), F+ RNA coliphages isolated from human waste were distinctively separate from those isolated from other animal sources. However, F+ DNA coliphages were not significantly different or separate in the PCoA. These results demonstrate that proper analysis of F+ RNA coliphages can effectively distinguish fecal sources.

Differential arsenic mobilization from As-bearing ferrihydrite by iron-respiring Shewanella strains with different arsenic-reducing activities.

Arsenic immobilization and release in the environment is significantly influenced by bacterial oxidation and reduction of arsenic and arsenic-bearing minerals. In this study, we tested three iron-reducing bacteria, Shewanella oneidensis MR-1, Shewanella sp. HN-41, and Shewanella putrefaciens 200, which have diverse arsenate-reducing activities with regard to reduction of an As-bearing ferrihydrite slurry. In the cultures of S. oneidensis MR-1 and Shewanella sp. HN-41, which are not capable of respiratory reduction of As(V) to As(III), arsenic was maintained predominantly in its pentavalent form, existing in particulate poorly crystalline As-bearing ferrihydrite and formed small quantities of a stable ferrous arsenate [Fe3(AsO4)2] precipitate. However, in the culture of the As(V) reducer, S. putrefaciens 200, As(V) was reduced to As(III) and a small fraction of As-bearing ferrihydrite was transformed into ribbon-shaped siderite that subsequently re-released arsenic into the liquid phase. Our results indicated that release of arsenic and formation of diverse secondary nanoscale Fe-As minerals are specifically closely related to the arsenic-reducing abilities of different bacteria. Therefore, bacterial arsenic reduction appears to significantly influence As mobilization in soils, minerals, and other Fe-rich environments.

Molecular cloning, expression and characterization of a glycosyltransferase from rice

Secondary plant metabolites undergo several modification reactions, including glycosylation. Glycosylation, which is mediated by UDP-glycosyltransferase (UGT), plays a role in the storage of secondary metabolites and in defending plants against stress. In this study, we cloned one of the glycosyltransferases from rice, RUGT-5 resulting in 40–42% sequence homology with UGTs from other plants. RUGT-5 was functionally expressed as a glutathione S-transferase fusion protein in Escherichia coli and was then purified. Eight different flavonoids were used as tentative substrates. HPLC profiling of reaction products displayed at least two peaks. Glycosylation positions were located at the hydroxyl groups at C-3, C-7 or C-4′ flavonoid positions. The most efficient substrate was kaempferol, followed by apigenin, genistein and luteolin, in that order. According to in vitro results and the composition of rice flavonoids the in vivo substrate of RUGT-5 was predicted to be kaempferol or apigenin. To our knowledge, this is the first time that the function of a rice UGT has been characterized.

Flavonoids biotransformation by bacterial non-heme dioxygenases, biphenyl and naphthalene dioxygenase

This review details recent progresses in the flavonoid biotransformation by bacterial non-heme dioxygenases, biphenyl dioxygenase (BDO), and naphthalene dioxygenase (NDO), which can initially activate biphenyl and naphthalene with insertion of dioxygen in stereospecfic and regiospecific manners. Flavone, isoflavone, flavanone, and isoflavanol were biotransformed by BDO from Pseudomonas pseudoalcaligenes KF707 and NDO from Pseudomonas sp. strain NCIB9816-4, respectively. In general, BDO showed wide range of substrate spectrum and produced the oxidized products, whereas NDO only metabolized flat two-dimensional substrates of flavone and isoflavone. Furthermore, biotransformation of B-ring skewed substrates, flavanone and isoflavanol, by BDO produced the epoxide products, instead of dihydrodiols. These results support the idea that substrate-driven reactivity alteration of the Fe-oxo active species may occur in the active site of non-heme dioxygenases. The study of flavonoid biotransformation by structurally-well defined BDO and NDO will provide the substrate structure and reactivity relationships and eventually establish the production of non-plant-originated flavonoids by means of microbial biotechnology.

Mercury capture into biogenic amorphous selenium nanospheres produced by mercury resistant Shewanella putrefaciens 200.

Shewanella putrefaciens 200, resistant to high concentration of Hg(II), was selected for co-removal of mercury and selenium from aqueous medium. Biogenic Hg(0) reduced from Hg(II) by S. putrefaciens 200 was captured into extracellular amorphous selenium nanospheres, resulting in the formation of stable HgSe nanoparticles. This bacterial reduction could be a new strategy for mercury removal from aquatic environments without secondary pollution of mercury methylation or Hg(0) volatilization.

Identification of fungal metabolites of anticonvulsant drug carbamazepine.

Carbamazepine, which has been used in the treatments of epilepsy, is often found in the environment. Although metabolism of carbamazepine by humans and rats has been characterized, the environmental fate of carbamazepine has not been studied. In this study, two model fungi Cunninghamella elegans ATCC 9245 and Umbelopsis ramanniana R-56, which have previously shown diverse metabolic activities, were tested for metabolism of carbamazepine. Both fungi produced three metabolites each (C1–C3 and M1–M3). All six metabolites showed [M + H]+ at m/z 253, suggesting addition of one oxygen to the parent compound. High-performance liquid chromatography and liquid chromatography–mass spectrometric analysis detected 10, 11-dihydro-10, 11-epoxycarbamazepine as a major product (C3 (47%) and M3 (85%)) and 3-hydroxycarbamazepine (C2 (15%) and M2 (7%)) from carbamazepine through mixed mono-oxidation reactions in both fungal strains. C. elegans was confirmed to produce 2-hydroxycarbamazepine (C1 (38%)) while U. ramanniana produced a yet unidentified ring-hydroxylated metabolite (M1 (8%)). The current study suggests that carbamazepine is likely to be subjected to initially diverse mono-oxygenation reactions by fungal metabolisms, resulting in the formation of the corresponding metabolites, which were similarly found in mammalian metabolisms.