A novel electrochemically active and Fe(III)-reducing bacterium phylogenetically related to Aeromonas hydrophila,isolated from a microbial fuel cell

A facultative anaerobic bacterium was isolated from a mediator-less microbial fuel cell fed with artificial wastewater containing acetate and designated as PA3. The isolate was identified as a strain of Aeromonas hydrophila based on its biochemical, physiological and morphological characteristics as well as 16S rDNA sequence analysis and DNA-DNA hybridization. PA3 used glucose, glycerol, pyruvate and hydrogen to reduce Fe(III), nitrate and sulfate. Cyclic voltammetry showed that PA3 was electrochemically active and was the culture collection strain A. hydrophila KCTC 2358. Electricity was generated from a fuel cell-type reactor, the anode compartment of which was inoculated with cell suspensions of the isolate or A. hydrophila KCTC 2358. The electrochemical activities are novel characteristics of A. hydrophila.     

Dissimilatory Fe(III) reduction by an electrochemically active lactic acid bacterium phylogenetically related to Enterococcus gallinarum isolated from submerged soil

AIMS: The isolation and identification of a glucose-oxidizing Fe(III)-reducing bacteria (FRB) with electrochemical activity from an anoxic environment, and characterization of the role of Fe(III) in its metabolism. METHODS AND RESULTS: A Gram-positive (Firmicutes), nonmotile, coccoid and facultative anaerobic FRB was isolated based on its ability to reduce Fe(III). Using the Vitek Gram-positive identification card kit and 16S rRNA gene sequence analysis, the isolate was identified as Enterococcus gallinarum, designated strain MG25. On glucose this isolate produced lactate plus small amounts of acetate, formate and CO2 and its growth rates were similar in the presence and absence of Fe(O)OH. These results suggest that MG25 can couple glucose oxidation to Fe(III) reduction, but without conservation of energy to support growth. Cyclic voltammetry showed that strain MG25 was electrochemically active. CONCLUSIONS: An electrochemically active and FRB, E. gallinarum MG25, was isolated from submerged soil. Fe(III) is used in the bacterial metabolism as an electron sink. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report concerning the electrochemical activity of glucose-oxidizing FRB, E. gallinarum. This organism and others like it could be used as new biocatalysts to improve the performance of a mediator-less microbial fuel cell.     

Bacterial community structure, compartmentalization and activity in a microbial fuel cell

AIMS: To characterize bacterial populations and their activities within a microbial fuel cell (MFC), using cultivation-independent and cultivation approaches. METHODS AND RESULTS: Electron microscopic observations showed that the fuel cell electrode had a microbial biofilm attached to its surface with loosely associated microbial clumps. Bacterial 16S rRNA gene libraries were constructed and analysed from each of four compartments within the fuel cell: the planktonic community; the membrane biofilm; bacterial clumps (BC) and the anode biofilm. Results showed that the bacterial community structure varied significantly between these compartments. It was observed that Gammaproteobacteria phylotypes were present at higher numbers within libraries from the BC and electrode biofilm compared with other parts of the fuel cell. Community structure of the MFC determined by analyses of bacterial 16S rRNA gene libraries and anaerobic cultivation showed excellent agreement with community profiles from denaturing gradient gel electrophoresis (DGGE) analysis. CONCLUSIONS: Members of the family Enterobacteriaceae, such as Klebsiella sp. and Enterobacter sp. and other Gammaproteobacteria with Fe(III)-reducing and electrochemical activity had a significant potential for energy generation in this system. SIGNIFICANCE AND IMPACT OF THE STUDY: This study has shown that electrochemically active bacteria can be enriched using an electrochemical fuel cell.     

New exoelectrogen Citrobacter sp. SX-1 isolated from a microbial fuel cell

Aims: Isolation, identification and characterization of a new exoelectrogenic bacterium from a microbial fuel cell (MFC). Methods and Results: Exoelectrogenic bacterial strain SX‐1 was isolated from a mediator‐less MFC by conventional plating techniques with ferric citrate as electron acceptor under anaerobic condition. Phylogenetic analysis of the 16S rDNA sequence revealed that it was related to the members of Citrobacter genus with Citrobacter sp. sdy‐48 being the most closely related species. The bacterial strain SX‐1 produced electricity from citrate, acetate, glucose, sucrose, glycerol and lactose in MFCs with the highest current density of 205 mA m^?2 generated from citrate. Cyclic voltammetry analysis indicated that membrane‐associated proteins may play an important role in facilitating the electrons transferring from bacteria to electrode. Conclusions: This is the first study that demonstrates that Citrobacter species can transfer electrons to extracellular electron acceptors. Citrobacter strain SX‐1 is capable of generating electricity from a wide range of substrates in MFCs. Significance and Impact of the Study: This finding increases the known diversity of power generating exoelectrogens and provided a new strain to explore the mechanisms of extracellular electron transfer from bacteria to electrode. The wide range of substrate utilization by SX‐1 increases the application potential of MFCs in renewable energy generation and waste treatment.     

From microbial fuel cell (MFC) to microbial electrochemical snorkel (MES): maximizing chemical oxygen demand (COD) removal from wastewater

The paper introduces the concept of the microbial electrochemical snorkel (MES), a simplified design of a “short-circuited” microbial fuel cell (MFC). The MES cannot provide current but it is optimized for wastewater treatment. An electrochemically active biofilm (EAB) was grown on graphite felt under constant polarization in an urban wastewater. Controlling the electrode potential and inoculating the bioreactor with a suspension of an established EAB improved the performance and the reproducibility of the anodes. Anodes, colonized by an EAB were tested for the chemical oxygen demand (COD) removal from urban wastewater using a variety of bio-electrochemical processes (microbial electrolysis, MFC, MES). The MES technology, as well as a short-circuited MFC, led to a COD removal 57% higher than a 1000 Ω-connected MFC, confirming the potential for wastewater treatment.     

Antibacterial activity of a disaccharide isolated from Streptomyces sp. strain JJ45 against Xanthomonas sp.

Of the 316 actinomycetes strains isolated from various habitats, Streptomyces sp. strain JJ45 showed the strongest antibiotic activity against the plant pathogenic bacteria Xanthomonas campestris pv. campestris and was thus chosen for further study. The 16S rRNA gene sequence (1500 bp) and rpoB gene partial sequence (306 bp) of Streptomyces strains JJ45A and JJ45B were determined. The respective strain JJ45B sequences exhibited 96.8% identity with the Streptococcus gelaticus 16S rRNA gene sequence and 98.4% identity with the Streptococcus vinaceus ATCC 27478 rpoB partial sequence. The fermentation broth of the JJ45B strain was extracted to find an inhibitor of bacterial growth. The distilled water extract showed the highest activity against pathogenic bacteria. The active molecule was isolated by column chromatography on polyacrylamide or silica gel, thin-layer chromatography, and HPLC. It showed growth inhibition activity only toward phytopathogenic Xanthomonas sp. The structure of the compound was identified as α- l -sorbofuranose (3→2)-β- d -altrofuranose based on the interpretation of the nuclear magnetic resonance spectra.     

Photoluminescence imaging of europium (III)‐doped γ‐Al2O3 nanofiber structures

Aluminium oxide (Al₂O₃) has widely been used for catalysts, insulators, and composite materials for diverse applications. Herein, we demonstrated if γ‐Al₂O₃ was useful as a luminescence support material for europium (Eu) (III) activator ion. The hydrothermal method and post‐thermal treatment at 800°C were employed to synthesize Eu(III)‐doped γ‐Al₂O₃ nanofibre structures. Luminescence characteristics of Eu(III) ions in Al₂O₃ matrix were fully understood by taking 2D and 3D‐photoluminescence imaging profiles. Various sharp emissions between 580 to 720 nm were assigned to the ⁵D₀→⁷F_J (J = 0, 1, 2, 3, 4) transitions of Eu(III) activators. On the basis of X‐ray diffraction crystallography, Auger elemental mapping and the asymmetry ratio, Eu(III) ions were found to be well doped into the γ‐Al₂O₃ matrix at a low (1 mol%) doping level. A broad emission at 460 nm was substantially increased upon higher (2 mol%) Eu(III) doping due to defect creation. The first 3D photoluminescence imaging profiles highlight detailed understanding of emission characteristics of Eu(III) ions in Al oxide‐based phosphor materials and their potential applications.     

Crystal structure of the apo form of a β‐transaminase from Mesorhizobium sp. strain LUK

Pyridoxal 5′‐phosphate (PLP)‐dependent β‐transaminases (βTAs) reversibly catalyze transamination reactions by recognizing amino groups linked to the β‐carbon atoms of their substrates. Although several βTA structures have been determined as holo forms containing PLP, little is known about the effect of PLP on the conversion of the apo structure to the holo structure. We determined the crystal structure of the apo form of a βTA from Mesorhizobium sp. strain LUK at 2.2 Å resolution to elucidate how PLP affects the βTA structure. The structure revealed three major disordered regions near the active site. Structural comparison with the holo form also showed that the disordered regions in the apo form are ordered and partially adopt secondary structures in the holo form. These findings suggest that PLP incorporation into the active site contributes to the structural stability of the active site architecture, thereby forming the complete active site. Our results provide novel structural insights into the role of PLP in terms of active site formation.     

Electricity generation from the treatment of wastewater with a hybrid up‐flow microbial fuel cell

The performance of a prototype up‐flow single‐chambered microbial fuel cell (MFC) for electrical power generation using brewery wastewater as fuel is reported. The designed reactor consisted of three zones, namely a lower anaerobic digestion zone, a central MFC zone, and an upper effluent clarifier zone. Tests were conducted in batch mode using a beer wastewater as the fuel/electron donor (COD concentration: 430 mg/L) and mixed consortia (both sewage microflora and anaerobic sludge) as a source of electrogenic bacteria. A stable current density of ～2,270 mA/m² was generated under continuous polarization with a constant external resistance (0.01 kΩ) and cell polarization gave a peak power density of 330 mW/m² at a current density of 1,680 mA/m². Electrochemical impedance analysis showed that the overall internal resistance of the reactor was quite low, that is, 8.0 Ω. Cyclic voltammetric analysis of the anodic biofilm at low scan rate revealed quite complex processes at the anode, with three redox peaks, at potentials of 116, 214, and 319 mV (vs. NHE). Biotechnol. Bioeng. 2010;107: 52–58. © 2010 Wiley Periodicals, Inc.     

Characterization of α‐rhamnosidase activity from a Patagonian Pichia guilliermondii wine strain

Aims: The purpose of this study was to characterize the α‐l ‐rhamnosidase of Pichia guilliermondii NPCC1053 indigenous wine strain from North‐Patagonian region. Methods and Results: The optimization of yeast culture conditions was carried out and the effects of oenological parameters on α‐l ‐rhamnosidase activity were evaluated. Additionally, the effect of direct contact with must and wine on α‐l ‐rhamnosidase activity was assayed. This strain showed an intracellular inducible α‐l ‐rhamnosidase activity. This enzyme was active at pH, glucose and SO₂ concentrations usually found at the beginning of the fermentation as well as retained high levels of activity after 24 h of incubation in must. Furthermore, P. guilliermondiiα‐l ‐rhamnosidase was able to release monoterpenols and alcohols from grape glycosidic extracts. Conclusions: The α‐l ‐rhamnosidase belonging to P. guilliermondii indigenous wine yeast strain showed mainly an intracellular location and evidenced interesting oenological characteristics. Significance and Impact of the Study: This study contributes to the knowledge of α‐l ‐rhamnosidases from yeast origin because at present, there are few reports about this enzymatic activity in these micro‐organisms. In addition, this work is relevant to the regional wine industry considering that this enzyme could be used in the production of more aromatic young wines.     

Cytotoxic (9βH)‐Pimarane and (9βH)‐17‐Norpimarane Diterpenes from the Tuber of Icacina trichantha

Three (9βH)‐pimaranes, 1, 2 , and 3 , and two (9βH)‐17‐norpimaranes, 4 and 5 , belonging to a rare compound class in nature, were obtained from the tubers of Icacina trichantha for the first time. Compound 1 is a new natural product, and 2 – 5 have been previously reported. The structures were elucidated based on NMR and MS data, and optical rotation values. The absolute configurations of (9βH)‐pimaranes were unambiguously established based on X‐ray crystallographic analysis. Full NMR signal assignments for the known compounds 2, 4 , and 5 , which were not available in previous publications, are also reported. All five isolates displayed cytotoxic activities on MDA‐MB‐435 cells (IC₅₀ 0.66–6.44 μM ), while 2, 3 , and 4 also exhibited cytotoxicities on HT‐29 cells (IC₅₀ 3.00–4.94 μM ).     

β‐arrestin2/miR‐155/GSK3β regulates transition of 5′‐azacytizine‐induced Sca‐1‐positive cells to cardiomyocytes

Stem‐cell antigen 1–positive (Sca‐1+) cardiac stem cells (CSCs), a vital kind of CSCs in humans, promote cardiac repair in vivo and can differentiate to cardiomyocytes with 5′‐azacytizine treatment in vitro. However, the underlying molecular mechanisms are unknown. β‐arrestin2 is an important scaffold protein and highly expressed in the heart. To explore the function of β‐arrestin2 in Sca‐1+ CSC differentiation, we used β‐arrestin2–knockout mice and overexpression strategies. Real‐time PCR revealed that β‐arrestin2 promoted 5′‐azacytizine‐induced Sca‐1+ CSC differentiation in vitro. Because the microRNA 155 (miR‐155) may regulate β‐arrestin2 expression, we detected its role and relationship with β‐arrestin2 and glycogen synthase kinase 3 (GSK3β), another probable target of miR‐155. Real‐time PCR revealed that miR‐155, inhibited by β‐arrestin2, impaired 5′‐azacytizine‐induced Sca‐1+ CSC differentiation. On luciferase report assay, miR‐155 could inhibit the activity of β‐arrestin2 and GSK3β, which suggests a loop pathway between miR‐155 and β‐arrestin2. Furthermore, β‐arrestin2‐knockout inhibited the activity of GSK3β. Akt, the upstream inhibitor of GSK3β, was inhibited in β‐arrestin2‐Knockout mice, so the activity of GSK3β was regulated by β‐arrestin2 not Akt. We transplanted Sca‐1+ CSCs from β‐arrestin2‐knockout mice to mice with myocardial infarction and found similar protective functions as in wild‐type mice but impaired arterial elastance. Furthermore, low level of β‐arrestin2 agreed with decreased phosphorylation of AKT and increased phophorylation of GSK3β, similar to in vitro findings. The β‐arrestin2/miR‐155/GSK3β pathway may be a new mechanism with implications for treatment of heart disease.     

A novel Fe(II)/α‐ketoglutarate‐dependent dioxygenase from Burkholderia ambifaria has β‐hydroxylating activity of N‐succinyl l‐leucine

An Fe(II)/α‐ketoglutarate‐dependent dioxygenase, SadA, was obtained from Burkholderia ambifaria AMMD and heterologously expressed in Escherichia coli. Purified recombinant SadA had catalytic activity towards several N‐substituted l‐amino acids, which was especially strong with N‐succinyl l‐leucine. With the NMR and LC‐MS analysis, SadA converted N‐succinyl l‐leucine into N‐succinyl l‐threo‐β‐hydroxyleucine with >99% diastereoselectivity. SadA is the first enzyme catalysing β‐hydroxylation of aliphatic amino acid‐related substances and a potent biocatalyst for the preparation of optically active β‐hydroxy amino acids.     

The cleidocranial dysplasia‐related R131G mutation in the Runt‐related transcription factor RUNX2 disrupts binding to DNA but not CBF‐β

Cleidocranial dysplasia (CCD) is caused by haploinsufficiency in RUNX2 function. We have previously identified a series of RUNX2 mutations in Korean CCD patients, including a novel R131G missense mutation in the Runt‐homology domain. Here, we examine the functional consequences of the RUNX2^R131G mutation, which could potentially affect DNA binding, nuclear localization signal, and/or heterodimerization with core‐binding factor‐β (CBF‐β). Immunofluorescence microscopy and western blot analysis with subcellular fractions show that RUNX2^R131G is localized in the nucleus. Immunoprecipitation analysis reveals that heterodimerization with CBF‐β is retained. However, precipitation assays with biotinylated oligonucleotides and reporter gene assays with RUNX2 responsive promoters together reveal that DNA‐binding activity and consequently the transactivation of potential of RUNX2^R131G is abrogated. We conclude that loss of DNA binding, but not nuclear localization or CBF‐β heterodimerization, causes RUNX2 haploinsufficiency in patients with the RUNX2^R131G mutation. Retention of specific functions including nuclear localization and binding to CBF‐β of the RUNX2^R131G mutation may render the mutant protein an effective competitor that interferes with wild‐type function. J. Cell. Biochem. 110: 97–103, 2010. © 2010 Wiley‐Liss, Inc.