PmoB subunit of particulate methane monooxygenase (pMMO) in Methylococcus capsulatus (Bath): The Cu^I sponge and its function

In this study, we describe efforts to clarify the role of the copper cofactors associated with subunit B (PmoB) of the particulate methane monooxygenase (pMMO) from Methylococcus capsulatus (Bath) (M. capsulatus). This subunit exhibits strong affinity toward Cu^I ions. To elucidate the high copper affinity of the subunit, the full-length PmoB, and the N-terminal truncated mutants PmoB_(33–414) and PmoB_(55–414), each fused to the maltose-binding protein (MBP), are cloned and over-expressed into Escherichia coli (E. coli) K12 TB1 cells. The Y374F, Y374S and M300L mutants of these protein constructs are also studied. When this E. coli is grown with the pmoB gene in 1.0 mM Cu^(II), it behaves like M. capsulatus (Bath) cultured under high copper stresswith abundant membrane accumulation and high CuI content. The recombinantPmoB proteins are verified by Western blotting of antibodies directed against the MBP sub-domain in each of the copper-enriched PmoB proteins. Cu K-edge X-rayabsorption near edge spectroscopy (XANES) of the copper ions confirms that all the PmoB recombinants are Cu^I proteins. All the PmoB proteins show evidence of a “dicopper site” according to analysis of the Cu extended X-ray absorption edge fine structure (EXAFS) of the membranes. No specific activities toward methane and propene oxidation are observed with the recombinant membrane-bound PmoB proteins. However, significant production of hydrogen peroxide is observed in the case of the PmoB_(33–414) mutant. Reaction of the dicopper site with dioxygenproduces hydrogen peroxide and leads to oxidation of the CuI ions residing in the C-terminal sub-domain of the PmoB subunit

PmoB subunit of particulate methane monooxygenase (pMMO) in Methylococcus capsulatus (Bath): The Cu^I sponge and its function

In this study, we describe efforts to clarify the role of the copper cofactors associated with subunit B (PmoB) of the particulate methane monooxygenase (pMMO) from Methylococcus capsulatus (Bath) (M. capsulatus). This subunit exhibits strong affinity toward Cu^I ions. To elucidate the high copper affinity of the subunit, the full-length PmoB, and the N-terminal truncated mutants PmoB_(33–414) and PmoB_(55–414), each fused to the maltose-binding protein (MBP), are cloned and over-expressed into Escherichia coli (E. coli) K12 TB1 cells. The Y374F, Y374S and M300L mutants of these protein constructs are also studied. When this E. coli is grown with the pmoB gene in 1.0 mM Cu^(II), it behaves like M. capsulatus (Bath) cultured under high copper stresswith abundant membrane accumulation and high CuI content. The recombinantPmoB proteins are verified by Western blotting of antibodies directed against the MBP sub-domain in each of the copper-enriched PmoB proteins. Cu K-edge X-rayabsorption near edge spectroscopy (XANES) of the copper ions confirms that all the PmoB recombinants are Cu^I proteins. All the PmoB proteins show evidence of a “dicopper site” according to analysis of the Cu extended X-ray absorption edge fine structure (EXAFS) of the membranes. No specific activities toward methane and propene oxidation are observed with the recombinant membrane-bound PmoB proteins. However, significant production of hydrogen peroxide is observed in the case of the PmoB_(33–414) mutant. Reaction of the dicopper site with dioxygenproduces hydrogen peroxide and leads to oxidation of the CuI ions residing in the C-terminal sub-domain of the PmoB subunit

An Overview of Regional Experiments on Biomass Burning Aerosols and Related Pollutants in Southeast Asia: From BASE-ASIA and the Dongsha Experiment to 7-SEAS

By modulating the Earth-atmosphere energy, hydrological and biogeochemical cycles, and affecting regional-to-global weather and climate, biomass burning is recognized as one of the major factors affecting the global carbon cycle. However, few comprehensive and wide-ranging experiments have been conducted to characterize biomass-burning pollutants in Southeast Asia (SEA) or assess their regional impact on meteorology, the hydrological cycle, the radiative budget, or climate change. Recently, BASEASIA (Biomass-burning Aerosols in South-East Asia: Smoke Impact Assessment) and the 7-SEAS (7- South-East Asian Studies) Dongsha Experiment were conducted during the spring seasons of 2006 and 2010 in northern SEA, respectively, to characterize the chemical, physical, and radiative properties of biomass-burning emissions near the source regions, and assess their effects. This paper provides an overview of results from these two campaigns and related studies collected in this special issue, entitled Observation, modeling and impact studies of biomass burning and pollution in the SE Asian Environment. This volume includes 28 papers, which provide a synopsis of the experiments, regional weatherclimate, chemical characterization of biomass-burning aerosols and related pollutants in source and sink regions, the spatial distribution of air toxics (atmospheric mercury and dioxins) in source and remote areas, a characterization of aerosol physical, optical, and radiative properties, as well as modeling and impact studies. These studies, taken together, provide the first relatively complete dataset of aerosol chemistry and physical observations conducted in the sourcesink region in the northern SEA, with particular emphasis on the marine boundary layer and lower free troposphere (LFT). The data, analysis and modeling included in these papers advance our present knowledge of source characterization of biomass-burning pollutants near the source regions as well as the physical and chemical processes along transport pathways. In addition, we raise key questions to be addressed by a coming deployment during springtime 2013 in northern SEA, named 7-SEASBASELInE (Biomass-burning Aerosols Stratocumulus Environment: Lifecycles and Interactions Experiment). This campaign will include a synergistic approach for further exploring many key atmospheric processes (e.g., complex aerosol-cloud interactions) and impacts of biomass burning on the surface-atmosphere energy budgets during the lifecycles of biomass burning emissions

Biosafety of Non-Surface Modified Carbon Nanocapsules as a Potential Alternative to Carbon Nanotubes for Drug Delivery Purposes

BACKGROUND: Carbon nanotubes (CNTs) have found wide success in circuitry, photovoltaics, and other applications. In contrast, several hurdles exist in using CNTs towards applications in drug delivery. Raw, non-modified CNTs are widely known for their toxicity. As such, many have attempted to reduce CNT toxicity for intravenous drug delivery purposes by post-process surface modification. Alternatively, a novel sphere-like carbon nanocapsule (CNC) developed by the arc-discharge method holds similar electric and thermal conductivities, as well as high strength. This study investigated the systemic toxicity and biocompatibility of different non-surface modified carbon nanomaterials in mice, including multi-walled carbon nanotubes (MWCNTs), single-walled carbon nanotubes (SWCNTs), carbon nanocapsules (CNCs), and C ₆₀ fullerene (C ₆₀). The retention of the nanomaterials and systemic effects after intravenous injections were studied. METHODOLOGY AND PRINCIPAL FINDINGS: MWCNTs, SWCNTs, CNCs, and C ₆₀ were injected intravenously into FVB mice and then sacrificed for tissue section examination. Inflammatory cytokine levels were evaluated with ELISA. Mice receiving injection of MWCNTs or SWCNTs at 50 µg/g b.w. died while C ₆₀ injected group survived at a 50% rate. Surprisingly, mortality rate of mice injected with CNCs was only at 10%. Tissue sections revealed that most carbon nanomaterials retained in the lung. Furthermore, serum and lung-tissue cytokine levels did not reveal any inflammatory response compared to those in mice receiving normal saline injection. CONCLUSION: Carbon nanocapsules are more biocompatible than other carbon nanomaterials and are more suitable for intravenous drug delivery. These results indicate potential biomedical use of non-surface modified carbon allotrope. Additionally, functionalization of the carbon nanocapsules could further enhance dispersion and biocompatibility for intravenous injection

Invasive fungal infection among hematopoietic stem cell transplantation patients with mechanical ventilation in the intensive care unit

<p>Abstract</p> <p>Background</p> <p>Invasive fungal infection (IFI) is associated with high morbidity and high mortality in hematopoietic stem cell transplantation (HSCT) patientsThe purpose of this study was to assess the characteristics and outcomes of HSCT patients with IFIs who are undergoing MV at a single institution in Taiwan.</p> <p>Methods</p> <p>We performed an observational retrospective analysis of IFIs in HSCT patients undergoing mechanical ventilation (MV) in an intensive care unit (ICU) from the year 2000 to 2009. The characteristics of these HSCT patients and risk factors related to IFIs were evaluated. The status of discharge, length of ICU stay, date of death and cause of death were also recorded.</p> <p>Results</p> <p>There were 326 HSCT patients at the Linkou Chang-Gung Memorial Hospital (Taipei, Taiwan) during the study period. Sixty of these patients (18%) were transferred to the ICU and placed on mechanical ventilators. A total of 20 of these 60 patients (33%) had IFIs. Multivariate analysis indicated that independent risk factors for IFI were admission to an ICU more than 40 days after HSCT, graft versus host disease (GVHD), and high dose corticosteroid (<it>p </it>< 0.01 for all). The overall ICU mortality rate was 88% (53 of 60 patients), and was not significantly different for patients with IFIs (85%) and those without IFIs (90%, <it>p </it>= 0.676).</p> <p>Conclusion</p> <p>There was a high incidence of IFIs in HSCT patients requiring MV in the ICU in our study cohort. The independent risk factors for IFI are ICU admission more than 40 days after HSCT, GVHD, and use of high-dose corticosteroid.</p

Toll-like receptor 2 gene polymorphisms, pulmonary tuberculosis, and natural killer cell counts

<p>Abstract</p> <p>Background</p> <p>To investigate whether the toll-like receptor 2 polymorphisms could influence susceptibility to pulmonary TB, its phenotypes, and blood lymphocyte subsets.</p> <p>Methods</p> <p>A total of 368 subjects, including 184 patients with pulmonary TB and 184 healthy controls, were examined for TLR2 polymorphisms over locus -100 (microsatellite guanine-thymine repeats), -16934 (T>A), -15607 (A>G), -196 to -174 (insertion>deletion), and 1350 (T>C). Eighty-six TB patients were examined to determine the peripheral blood lymphocyte subpopulations.</p> <p>Results</p> <p>We newly identified an association between the haplotype [A-G-(insertion)-T] and susceptibility to pulmonary TB (p = 0.006, false discovery rate q = 0.072). TB patients with systemic symptoms had a lower -196 to -174 deletion/deletion genotype frequency than those without systemic symptoms (5.7% vs. 17.7%; p = 0.01). TB patients with the deletion/deletion genotype had higher blood NK cell counts than those carrying the insertion allele (526 vs. 243.5 cells/μl, p = 0.009). TB patients with pleuritis had a higher 1350 CC genotype frequency than those without pleuritis (12.5% vs. 2.1%; p = 0.004). TB patients with the 1350 CC genotype had higher blood NK cell counts than those carrying the T allele (641 vs. 250 cells/μl, p = 0.004). TB patients carrying homozygous short alleles for GT repeats had higher blood NK cell counts than those carrying one or no short allele (641 vs. 250 cells/μl, p = 0.004).</p> <p>Conclusions</p> <p>TLR2 genetic polymorphisms influence susceptibility to pulmonary TB. TLR2 variants play a role in the development of TB phenotypes, probably by controlling the expansion of NK cells.</p

The genetic heterogeneity and mutational burden of engineered melanomas in zebrafish models.

BACKGROUND: Melanoma is the most deadly form of skin cancer. Expression of oncogenic BRAF or NRAS, which are frequently mutated in human melanomas, promote the formation of nevi but are not sufficient for tumorigenesis. Even with germline mutated p53, these engineered melanomas present with variable onset and pathology, implicating additional somatic mutations in a multi-hit tumorigenic process. RESULTS: To decipher the genetics of these melanomas, we sequence the protein coding exons of 53 primary melanomas generated from several BRAF(V600E) or NRAS(Q61K) driven transgenic zebrafish lines. We find that engineered zebrafish melanomas show an overall low mutation burden, which has a strong, inverse association with the number of initiating germline drivers. Although tumors reveal distinct mutation spectrums, they show mostly C > T transitions without UV light exposure, and enrichment of mutations in melanogenesis, p53 and MAPK signaling. Importantly, a recurrent amplification occurring with pre-configured drivers BRAF(V600E) and p53-/- suggests a novel path of BRAF cooperativity through the protein kinase A pathway. CONCLUSION: This is the first analysis of a melanoma mutational landscape in the absence of UV light, where tumors manifest with remarkably low mutation burden and high heterogeneity. Genotype specific amplification of protein kinase A in cooperation with BRAF and p53 mutation suggests the involvement of melanogenesis in these tumors. This work is important for defining the spectrum of events in BRAF or NRAS driven melanoma in the absence of UV light, and for informed exploitation of models such as transgenic zebrafish to better understand mechanisms leading to human melanoma formation