Flexibility in the receptor-binding domain of the enzymatic colicin E9 is required for toxicity against Escherichia coli cells

The events that occur after the binding of the enzymatic E colicins to Escherichia coli BtuB receptors that lead to translocation of the cytotoxic domain into the periplasmic space and, ultimately, cell killing are poorly understood. It has been suggested that unfolding of the coiled-coil Mull receptor binding domain of the E colicins may be an essential step that leads to the loss of immunity protein from the colicin and immunity protein complex and then triggers the events of translocation. We introduced pairs of cysteine mutations into the receptor binding domain of colicin E9 (ColE9) that resulted in the formation of a disulfide bond located near the middle or the top of the R domain. After dithiothreitol reduction, the ColE9 protein with the mutations L359C and F412C (ColE9 L359C-F412C) and the ColE9 protein with the mutations Y324C and L447C (ColE9 Y324C-L447C) were slightly less active than equivalent concentrations of ColE9. On oxidation with diamide, no significant biological activity was seen with the ColE9 L359C-F412C and the ColE9 Y324C-L447C mutant proteins; however diamide had no effect on the activity of ColE9. The presence of a disulfide bond was confirmed in both of the oxidized, mutant proteins by matrix-assisted laser desorption ionization-time of flight mass spectrometry. The loss of biological activity of the disulfide-containing mutant proteins was not due to an indirect effect on the properties of the translocation or DNase domains of the mutant colicins. The data are consistent with a requirement for the flexibility of the coiled-coil R domain after binding to BtuB

Near-Roadway Air Pollution and Coronary Heart Disease: Burden of Disease and Potential Impact of a Greenhouse Gas Reduction Strategy in Southern California

Background: Several studies have estimated the burden of coronary heart disease (CHD) mortality from ambient regional particulate matter ≤ 2.5 μm (PM2.5). The burden of near-roadway air pollution (NRAP) generally has not been examined, despite evidence of a causal link with CHD. Objective: We investigated the CHD burden from NRAP and compared it with the PM2.5 burden in the California South Coast Air Basin for 2008 and under a compact urban growth greenhouse gas reduction scenario for 2035. Methods: We estimated the population attributable fraction and number of CHD events attributable to residential traffic density, proximity to a major road, elemental carbon (EC), and PM2.5 compared with the expected disease burden if the population were exposed to background levels of air pollution. Results: In 2008, an estimated 1,300 CHD deaths (6.8% of the total) were attributable to traffic density, 430 deaths (2.4%) to residential proximity to a major road, and 690 (3.7%) to EC. There were 1,900 deaths (10.4%) attributable to PM2.5. Although reduced exposures in 2035 should result in smaller fractions of CHD attributable to traffic density, EC, and PM2.5, the numbers of estimated deaths attributable to each of these exposures are anticipated to increase to 2,500, 900, and 2,900, respectively, due to population aging. A similar pattern of increasing NRAP-attributable CHD hospitalizations was estimated to occur between 2008 and 2035. Conclusion: These results suggest that a large burden of preventable CHD mortality is attributable to NRAP and is likely to increase even with decreasing exposure by 2035 due to vulnerability of an aging population. Greenhouse gas reduction strategies developed to mitigate climate change offer unexploited opportunities for air pollution health co-benefits

Temporal and Spatial Patterns of Ambient Endotoxin Concentrations in Fresno, California

BackgroundEndotoxins are found in indoor dust generated by human activity and pets, in soil, and adsorbed onto the surfaces of ambient combustion particles. Endotoxin concentrations have been associated with respiratory symptoms and the risk of atopy and asthma in children.ObjectiveWe characterized the temporal and spatial variability of ambient endotoxin in Fresno/Clovis, California, located in California's Central Valley, to identify correlates and potential predictors of ambient endotoxin concentrations in a cohort of children with asthma [Fresno Asthmatic Children's Environment Study (FACES)].MethodsBetween May 2001 and October 2004, daily ambient endotoxin and air pollutants were collected at the central ambient monitoring site of the California Air Resources Board in Fresno and, for shorter time periods, at 10 schools and indoors and outdoors at 84 residences in the community. Analyses were restricted to May-October, the dry months during which endotoxin concentrations are highest.ResultsDaily endotoxin concentration patterns were determined mainly by meteorologic factors, particularly the degree of air stagnation. Overall concentrations were lowest in areas distant from agricultural activities. Highest concentrations were found in areas immediately downwind from agricultural/pasture land. Among three other measured air pollutants [fine particulate matter, elemental carbon (a marker of traffic in Fresno), and coarse particulate matter (PMc)], PMc was the only pollutant correlated with endotoxin. Endotoxin, however, was the most spatially variable.ConclusionsOur data support the need to evaluate the spatial/temporal variability of endotoxin concentrations, rather than relying on a few measurements made at one location, in studies of exposure and and respiratory health effects, particularly in children with asthma and other chronic respiratory diseases

Electronic structures and photophysics of d(8)-d(8) complexes

This work was supported by the NSF CCI Solar Fuels Program (CHE-1305124), the Arnold and Mabel Beckman Foundation, the Ministry of Education of the Czech Republic - grant LH13015, and the COST Action CM1405

Conditions Leading to Elevated PM_2.5 at Near-Road Monitoring Sites: Case Studies in Denver and Indianapolis

We examined two near-road monitoring sites where the daily PM2.5 readings were among the highest of any near-road monitoring location in the U.S. during 2014−2016: Denver, Colorado, in February 2014 and Indianapolis, Indiana, in November 2016. At the Denver site, which had the highest measured U.S. near-road 24-hr PM2.5 concentrations in 2014, concentrations exceeded the daily National Ambient Air Quality Standards (NAAQS) on three days during one week in 2014; the Indianapolis site had the second-highest number of daily exceedances of any near-road site in 2016 and the highest 3-year average PM2.5 of any near-road site during 2014−2016. Both sites had hourly pollutant, meteorological, and traffic data available, making them ideal for case studies. For both locations, we compared air pollution observations at the near-road site to observations at other sites in the urban area to calculate the near-road PM2.5 “increment” and evaluated the effects of changes in meteorology and traffic. The Denver near-road site consistently had the highest PM2.5 values in the Denver area, and was typically highest when winds were near-downwind, rather than directly downwind, to the freeway. Complex Denver site conditions (near-road buildings and roadway alignment) likely contributed to higher PM2.5 concentrations. The increment at Indianapolis was also highest under near-downwind, rather than directly downwind, conditions. At both sites, while the near-road site often had higher PM2.5 concentrations than nearby sites, there was no clear correlation between traffic conditions (vehicle speed, fleet mix) and the high PM2.5 concentrations

Rapid Detection of Colicin E9-Induced DNA Damage Using Escherichia coli Cells Carrying SOS Promoter-lux Fusions

ColE9 is a plasmid-encoded protein antibiotic produced by Escherichia coli and closely related species that kills E. coli cells expressing the BtuB receptor. The 15-kDa cytotoxic DNase domain of colicin E9 preferentially nicks double-stranded DNA at thymine bases and shares a common active-site structural motif with a variety of other nucleases, including the H-N-H homing endonucleases and the apoptotic CAD proteins of eukaryotes. Studies of the mechanism by which the DNase domain of ColE9 reaches the cytoplasm of E. coli cells are limited by the lack of a rapid, sensitive assay for the DNA damage that results. Here, we report the development of an SOS promoter-lux fusion reporter system for monitoring DNA damage in colicin-treated cells and illustrate the value of this reporter system in experiments that probe the mechanism and time required for the DNase domain of colicin E9 to reach the cytoplasm

Temporal and spatial patterns of ambient endotoxin concentrations in Fresno, California.

BackgroundEndotoxins are found in indoor dust generated by human activity and pets, in soil, and adsorbed onto the surfaces of ambient combustion particles. Endotoxin concentrations have been associated with respiratory symptoms and the risk of atopy and asthma in children.ObjectiveWe characterized the temporal and spatial variability of ambient endotoxin in Fresno/Clovis, California, located in California's Central Valley, to identify correlates and potential predictors of ambient endotoxin concentrations in a cohort of children with asthma [Fresno Asthmatic Children's Environment Study (FACES)].MethodsBetween May 2001 and October 2004, daily ambient endotoxin and air pollutants were collected at the central ambient monitoring site of the California Air Resources Board in Fresno and, for shorter time periods, at 10 schools and indoors and outdoors at 84 residences in the community. Analyses were restricted to May-October, the dry months during which endotoxin concentrations are highest.ResultsDaily endotoxin concentration patterns were determined mainly by meteorologic factors, particularly the degree of air stagnation. Overall concentrations were lowest in areas distant from agricultural activities. Highest concentrations were found in areas immediately downwind from agricultural/pasture land. Among three other measured air pollutants [fine particulate matter, elemental carbon (a marker of traffic in Fresno), and coarse particulate matter (PMc)], PMc was the only pollutant correlated with endotoxin. Endotoxin, however, was the most spatially variable.ConclusionsOur data support the need to evaluate the spatial/temporal variability of endotoxin concentrations, rather than relying on a few measurements made at one location, in studies of exposure and and respiratory health effects, particularly in children with asthma and other chronic respiratory diseases

Costs of coronary heart disease and mortality associated with near-roadway air pollution

Emerging evidence indicates that the near-roadway air pollution (NRAP) mixture contributes to CHD, yet few studies have evaluated the associated costs.; We integrated an assessment of NRAP-attributable CHD in Southern California with new methods to value the associated mortality and hospitalizations.; Based on population-weighted residential exposure to NRAP (traffic density, proximity to a major roadway and elemental carbon), we estimated the inflation-adjusted value of NRAP-attributable mortality and costs of hospitalizations that occurred in 2008. We also estimated anticipated costs in 2035 based on projected changes in population and in NRAP exposure associated with California's plans to reduce greenhouse gas emissions. For comparison, we estimated the value of CHD mortality attributable to PM less than 2.5μm in diameter (PM2.5) in both 2008 and 2035.; The value of CHD mortality attributable to NRAP in 2008 was between $3.8 and$11.5 billion, 23% (major roadway proximity) to 68% (traffic density) of the $16.8 billion attributable to regulated regional PM2.5. NRAP-attributable costs were projected to increase to$10.6 to $22 billion in 2035, depending on the NRAP metric. Cost of NRAP-attributable hospitalizations for CHD in 2008 was$48.6 million and was projected to increase to $51.4 million in 2035.; We developed an economic framework that can be used to estimate the benefits of regulations to improve air quality. CHD attributable to NRAP has a large economic impact that is expected to increase by 2035, largely due to an aging population. PM2.5-attributable costs may underestimate total value of air pollution-attributable CHD