Mononuclear copper active-oxygen complexes

There have been significant advances in the understanding of the dioxygen-activation chemistry of mononuclear copper monooxygenases such as peptidylglycine alpha-amidating monooxygenase and dopamine beta-monooxygenase (DbetaM). Recent structural and spectroscopic studies on a series of biomimetic model compounds have provided new and valuable insights into the key reactive intermediates involved in the dioxygen processing at the mononuclear copper reaction centers in biological systems.     

Lessons from comparative analysis of species-specific imprinted genes

Genomic imprinting is generally believed to be conserved in all mammals except for egg-laying monotremes, suggesting that it is closely related to placental and fetal growth. As expected, the imprinting status of most imprinted genes is conserved between mouse and human, and some are imprinted even in marsupials. On the other hand, a small number of genes were reported to exhibit species-specific imprinting that is not necessarily accounted for by either the placenta or conflict hypotheses. Since mouse and human represent a single, phylogenetically restricted clade in the mammalian class, a much broader comparison including mammals diverged earlier than rodents is necessary to fully understand the species-specificity and variation in evolution of genomic imprinting. Indeed, comparative analysis of a species-specific imprinted gene Impact using a broader range of mammals led us to propose an alternative dosage control hypothesis for the evolution of genomic imprinting.     

Isolation and characterization of novel denitrifying alkalithermophiles, AT-1 and AT-2

Two novel denitrifying alkalithermophilic bacteria, AT-1 and AT-2, were isolated from manure-amended soil. The isolates grew at 35–65°C with an optimum temperature at 50–60°C, and pH 6.5–10.0 with an optimum pH at 9.5. Both isolates were Gram-positive, facultative anaerobic, non-motile rod-shaped bacteria. A phylogenetic analysis based on 16S rRNA sequence data indicated that both AT-1 and AT-2 are members of the genus Anoxybacillus. DNA-DNA hybridization revealed moderate relatedness between AT-1 and AT-2 and one phylogenetically related strain, A. pushchinensis K1 (69.5 and 69.1%, respectively). Comparative analysis of morphology and biochemical characteristics of the two isolates also showed similarity to A. pushchinensis K1. Based on these results, we identified AT-1 and AT-2 as A. pushchinensis. To our knowledge, this is the first report of denitrifying bacterium isolated from alkalithermophilic Anoxybacillus spp.     

A case of intramuscular cysticercosis diagnosed definitively by mitochondrial DNA analysis of extremely calcified cysts

A case of obsolete intramuscular cysticercosis diagnosed definitively by mitochondrial DNA analysis of extremely calcified cysts was reported. X-ray and computed tomography findings highly suggested cysticercosis due to Taenia solium; however, no direct evidence of cysticercosis was obtained through serological or histopathological examinations. Mitochondrial DNA analysis of a histopathological specimen confirmed the causative agent to be the Asian genotype of T. solium.     

A functional model for pMMO (particulate methane monooxygenase): Hydroxylation of alkanes with H2O2 catalyzed by beta-diketiminatocopper(II) complexes

The reaction of copper(II) complexes supported by a series of beta-diketiminate ligands ((R1,R2)L, [(Dipp)N-C(R(2))-C(R(1))-C(R(2))-N(Dipp)](-), Dipp=2,6-diisopropylphenyl; see ) and H(2)O(2) has been examined spectroscopically at a low temperature. The beta-diketiminatocopper(II) complexes with R(2)=H (no substituent on the beta-carbon) provided a copper-oxygen intermediate that exhibited the same spectroscopic features as those of the bis(mu-oxo)dicopper(III) complex generated by the reaction of corresponding beta-diketiminatocopper(I) complex and O(2). On the other hand, the beta-diketiminatocopper(II) complexes with methyl substituent on the beta-carbon (R(2)=Me) did not produce such an intermediate in the same reaction. The beta-diketiminatocopper(II) complexes carrying an electron-withdrawing substituent on the alpha-carbon (R(1)=NO(2) or CN) but no beta-substituent (R(2)=H) exhibited a high catalytic activity in the oxygenation reaction of alkanes with H(2)O(2). Mechanism of the catalytic oxygenation reaction as well as the substituent effects of the ligands on the copper(II)-H(2)O(2) reactivity is discussed.     

Microbial community of a mesophilic propionate-degrading methanogenic consortium in chemostat cultivation analyzed based on 16S rRNA and acetate kinase genes

Abstract We constructed a mesophilic anaerobic chemostat that was continuously fed with synthetic wastewater containing propionate as the sole source of carbon and energy. Steady-state conditions were achieved below the critical dilution rate of 0.3 d ⁻¹ with almost complete substrate degradation. The propionate-degrading methanogenic communities in the chemostat at dilution rates of 0.01, 0.08, and 0.3 d ⁻¹ were analyzed using molecular biological techniques. Fluorescence in situ hybridization with archaeal and bacterial domain-specific probes showed that archaeal cells predominated throughout the three dilution rates. Archaeal-16S rRNA gene clone library analysis and quantitative real-time polymerase chain reaction studies showed that hydrogenotrophic methanogen rRNA genes closely related to Methanoculleus was detected at a dilution rate of 0.01 d ⁻¹ , whereas rRNA genes closely related to the Methanoculleus and Methanospirillum genera were detected at dilution rates of 0.08 and 0.3 d ⁻¹ . The aceticlastic methanogen, Methanosaeta , was detected throughout the three dilution rates. Bacterial-rRNA gene clone library analysis and denaturing gradient gel electrophoresis demonstrated that rRNA genes affiliated with the genus Syntrophobacter predominated at the low dilution rate, whereas rRNA genes affiliated with the phylum Firmicutes predominated at the higher dilution rates. A significant number of rRNA genes affiliated with the genus Pelotomaculum were detected at dilution rate of 0.3 d ⁻¹ . The diversity of genes encoding acetate kinase agreed closely with the results of the rRNA gene analysis. The dilution rates significantly altered the archaeal and bacterial communities in the propionate-fed chemostat.     

Mechanisms and pathophysiological implications of sinusoidal endothelial cell gap formation following treatment with galactosamine/endotoxin in mice

Neutrophil extravasation from sinusoids is a critical step for acute inflammatory tissue injury. However, the role of sinusoidal endothelial cells (SECs) in this process remains unclear. Matrix metalloproteinases (MMPs) have been shown to involve gap formation in SECs in several liver diseases. Therefore, the present study examined SEC modifications elicited by galactosamine (Gal)/endotoxin (ET). Treatment of male C3Heb/FeJ mice with Gal/ET or Gal/TNF caused the formation of numerous gaps in SECs at 4 h when no neutrophil extravasation occurred. Six hours after Gal/ET or Gal/TNF treatment, blood elements started to penetrate to the extrasinusoidal space through large gaps. Treatment with ET alone caused sinusoidal neutrophil accumulation but no gap formation, neutrophil extravasation, or hemorrhage. Gal/ET treatment increased hepatic MMP-2 and MMP-9 mRNA expression (6.7- and 11-fold, respectively). Pretreatment with 2-[(4-biphenylsulfonyl) amino]-3-phenyl-propionic acid, an MMP-2/MMP-9 inhibitor (5 mg/kg), minimized gap formation after Gal/ET and Gal/TNF treatment. The MMP inhibitor reduced injury only in the Gal/ET model mainly due to reduced TNF formation. The MMP inhibitor attenuated sinusoidal neutrophil accumulation at 6 h but failed to attenuate Gal/TNF-induced liver injury at 7 h due to excessive apoptosis. These results suggest that Gal/ET or Gal/TNF activates MMPs, which are responsible for SEC gap formation. Although the initial appearance of gap formation is independent of neutrophils, the gaps allow initial contact of neutrophils with damaged hepatocytes. In addition, MMP activation promotes neutrophil accumulation in sinusoids.     

Differential inhibitory effects of mu-opioids on substance P- and capsaicin-induced nociceptive behavior in mice

The antinociceptive mechanisms of the selective mu-opioid receptor agonists [D-Ala2,NMePhe4,Gly(ol)5]enkephalin (DAMGO), H-Tyr-D-Arg-Phe-beta-Ala-OH (TAPA) or H-Tyr-D-Arg-Phe-beta-Ala-NH2 (TAPA-NH2) against substance P (SP)- or capsaicin-elicited nociceptive behaviors was investigated in mice. DAMGO, TAPA or TAPA-NH2 given intrathecally inhibited the nociceptive behaviors elicited by intrathecally administered SP or capsaicin, and these antinociceptive effects were completely eliminated by intrathecal co-administration with D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2 (CTOP), a selective mu-opioid receptor antagonist. Pretreatment subcutaneously with naloxonazine, a selective mu1-opioid receptor antagonist, partially attenuated the antinociceptive effect of TAPA-NH2, but not DAMGO and TAPA, against SP. However, the antinociception induced by TAPA, but not DAMGO and TAPA-NH2, against capsaicin was significantly inhibited by naloxonazine. On the other hand, co-administration intrathecally with Tyr-D-Pro-Trp-Gly-NH2 (D-Pro2-Tyr-W-MIF-1), a selective mu2-opioid receptor antagonist, significantly attenuated the antinociceptive effects of DAMGO, but not TAPA and TAPA-NH2, against capsaicin, while the antinociceptions induced by three opioid peptides against SP were significantly inhibited by D-Pro2-Tyr-W-MIF-1. These results suggest that differential inhibitory mechanisms on pre- and postsynaptic sites in the spinal cord contribute to the antinociceptive effects of the three mu-opioid peptides.     

Identification of intestinal bacteria from Japanese flounder (Paralichthys olivaceus) and their ability to digest chitin

AIMS: The aim of the present study was to clarify the taxonomic status of intestinal bacteria isolated from Japanese flounder (Paralichthys olivaceus) and describe their ability to digest chitin. METHODS AND RESULTS: Phylogenetic analysis based on 16S ribosomal DNA (rDNA) sequences showed that 82 representative isolates were closely related to three major species of marine vibrios, Vibrio scophthalmi-Vibrio ichthyoenteri group (41 isolates), Vibrio fischeri (39 isolates) and Vibrio harveyi (two isolates), with similarities of 97.2-99.8%, 96.4-100% and 98.6-99.5% respectively. These findings indicate that V. scophthalmi-V. ichthyoenteri group is indigenous to the intestinal tract of Japanese flounder. Moreover, the ability of 82 isolates to digest chitin was examined using the agar plate method and PCR amplification of the chiA gene. The two V. harveyi isolates and 36 of 41 V. scophthalmi-V. ichthyoenteri isolates digested chitin and were chiA PCR positive, whereas all 39 V. fischeri isolates digested chitin but were chiA PCR negative. CONCLUSIONS: Intestinal bacteria from Japanese flounder were mainly composed of Vibrio scophthalmi-V. ichthyoenteri group and V. fischeri. Taken together, the results showed that 81 of 82 isolates could digest chitin. However, only 38 of these isolates possessed a chiA homologue which could be identified by PCR. SIGNIFICANCE AND IMPACT OF THE STUDY: The present study shows that Japanese flounder harbours bacteria of the V. scophthalmi-V. ichthyoenteri group, and these results are similar to what has been found for turbot (Scophthalmus maximus).