The structure of the substrate-free form of MurB, an essential enzyme for the synthesis of bacterial cell walls

AbstractBackground: The repeating disaccharide and pentapeptide units of the bacterial peptidoglycan layer are connected by a lactyl ether bridge biosynthesized from UDP-N-acetylglucosamine and phosphoenolpyruvate in sequential enol ether transfer and reduction steps catalyzed by MurA and MurB respectively. Knowledge of the structure and mechanism of the MurB enzyme will permit analysis of this unusual enol ether reduction reaction and may facilitate the design of inhibitors as candidate next-generation antimicrobial agents.Results The crystal structure of UDP-N-acetylenolpyruvylglucosamine reductase, MurB, has been solved at 3.0 å and compared with our previously reported structure of MurB complexed with its substrate enolpyruvyl-UDP-N-acetylglucosamine. Comparison of the liganded structure of MurB with this unliganded form reveals that the binding of substrate induces a substantial movement of domain 3 (residues 219–319) of the enzyme and a significant rearrangement of a loop within this domain. These ligand induced changes disrupt a stacking interaction between two tyrosines (Tyr190 and Tyr254) which lie at the side of the channel leading to the active site of the free enzyme.Conclusion The conformational change induced by enolpyruvyl-UDP-N-acetylglucosamine binding to MurB results in the closure of the substrate-binding channel over the substrate. Tyr190 swings over the channel opening and establishes a hydrogen bond with an oxygen of the α-phosphate of the sugar nucleotide substrate which is critical to substrate binding

The RhoA Guanine Nucleotide Exchange Factor, LARG, Mediates ICAM-1-Dependent Mechanotransduction in Endothelial Cells To Stimulate Transendothelial Migration

RhoA-mediated cytoskeletal rearrangements in endothelial cells (ECs) play an active role in leukocyte transendothelial cell migration (TEM), a normal physiological process in which leukocytes cross the endothelium to enter the underlying tissue. Although much has been learned about RhoA signaling pathways downstream from ICAM-1 in ECs, little is known about the consequences of the tractional forces that leukocytes generate on ECs as they migrate over the surface before TEM. We have found that after applying mechanical forces to ICAM-1 clusters, there is an increase in cellular stiffening and enhanced RhoA signaling compared with ICAM-1 clustering alone. We have identified that leukemia-associated Rho guanine nucleotide exchange factor (LARG), also known as Rho GEF 12 (ARHGEF12) acts downstream of clustered ICAM-1 to increase RhoA activity, and that this pathway is further enhanced by mechanical force on ICAM-1. Depletion of LARG decreases leukocyte crawling and inhibits TEM. To our knowledge, this is the first report of endothelial LARG regulating leukocyte behavior and EC stiffening in response to tractional forces generated by leukocytes

Features and Prognosis of Severe Malaria Caused by Plasmodium falciparum, Plasmodium vivax and Mixed Plasmodium Species in Papua New Guinean Children

BACKGROUND: Mortality from severe pediatric falciparum malaria appears low in Oceania but Plasmodium vivax is increasingly recognized as a cause of complications and death. The features and prognosis of mixed Plasmodium species infections are poorly characterized. Detailed prospective studies that include accurate malaria diagnosis and detection of co-morbidities are lacking. METHODS AND FINDINGS: We followed 340 Papua New Guinean (PNG) children with PCR-confirmed severe malaria (77.1% P. falciparum, 7.9% P. vivax, 14.7% P. falciparum/vivax) hospitalized over a 3-year period. Bacterial cultures were performed to identify co-incident sepsis. Clinical management was under national guidelines. Of 262 children with severe falciparum malaria, 30.9%, 24.8% and 23.2% had impaired consciousness, severe anemia, and metabolic acidosis/hyperlactatemia, respectively. Two (0.8%) presented with hypoglycemia, seven (2.7%) were discharged with neurologic impairment, and one child died (0.4%). The 27 severe vivax malaria cases presented with similar phenotypic features to the falciparum malaria cases but respiratory distress was five times more common (P=0.001); one child died (3.7%). The 50 children with P. falciparum/vivax infections shared phenotypic features of mono-species infections, but were more likely to present in deep coma and had the highest mortality (8.0%; P=0.003 vs falciparum malaria). Overall, bacterial cultures were positive in only two non-fatal cases. 83.6% of the children had alpha-thalassemia trait and seven with coma/impaired consciousness had South Asian ovalocytosis (SAO). CONCLUSIONS: The low mortality from severe falciparum malaria in PNG children may reflect protective genetic factors other than alpha-thalassemia trait/SAO, good nutrition, and/or infrequent co-incident sepsis. Severe vivax malaria had similar features but severe P. falciparum/vivax infections were associated with the most severe phenotype and worst prognosis

Tertiary-Treated Municipal Wastewater is a Significant Point Source of Antibiotic Resistance Genes Into Duluth-Superior Harbor

In this study, the impact of tertiary-treated municipal wastewater on the quantity of several antibiotic resistance determinants in Duluth-Superior Harbor was investigated by collecting surface water and sediment samples from 13 locations in Duluth-Superior Harbor, the St. Louis River, and Lake Superior. Quantitative PCR (qPCR) was used to target three different genes encoding resistance to tetracycline (tet(A), tet(X), and tet(W)), the gene encoding the integrase of class 1 integrons (intI1), and total bacterial abundance (16S rRNA genes) as well as total and human fecal contamination levels (16S rRNA genes specific to the genus Bacteroides). The quantities of tet(A), tet(X), tet(W), intI1, total Bacteroides, and human-specific Bacteroides were typically 20-fold higher in the tertiary-treated wastewater than in nearby surface water samples. In contrast, the quantities of these genes in the St. Louis River and Lake Superior were typically below detection. Analysis of sequences of tet(W) gene fragments from four different samples collected throughout the study site supported the conclusion that tertiary-treated municipal wastewater is a point source of resistance genes into Duluth-Superior Harbor. This study demonstrates that the discharge of exceptionally treated municipal wastewater can have a statistically significant effect on the quantities of antibiotic resistance genes in otherwise pristine surface waters

Antimicrobial Resistance among Campylobacter Strains, United States, 1997–2001

We summarize antimicrobial resistance surveillance data in human and chicken isolates of Campylobacter. Isolates were from a sentinel county study from 1989 through 1990 and from nine state health departments participating in National Antimicrobial Resistance Monitoring System for enteric bacteria (NARMS) from 1997 through 2001. None of the 297 C. jejuni or C. coli isolates tested from 1989 through 1990 was ciprofloxacin-resistant. From 1997 through 2001, a total of 1,553 human Campylobacter isolates were characterized: 1,471 (95%) were C. jejuni, 63 (4%) were C. coli, and 19 (1%) were other Campylobacter species. The prevalence of ciprofloxacin-resistant Campylobacter was 13% (28 of 217) in 1997 and 19% (75 of 384) in 2001; erythromycin resistance was 2% (4 of 217) in 1997 and 2% (8 of 384) in 2001. Ciprofloxacin-resistant Campylobacter was isolated from 10% of 180 chicken products purchased from grocery stores in three states in 1999. Ciprofloxacin resistance has emerged among Campylobacter since 1990 and has increased in prevalence since 1997

Modern optical astronomy: technology and impact of interferometry

The present `state of the art' and the path to future progress in high spatial resolution imaging interferometry is reviewed. The review begins with a treatment of the fundamentals of stellar optical interferometry, the origin, properties, optical effects of turbulence in the Earth's atmosphere, the passive methods that are applied on a single telescope to overcome atmospheric image degradation such as speckle interferometry, and various other techniques. These topics include differential speckle interferometry, speckle spectroscopy and polarimetry, phase diversity, wavefront shearing interferometry, phase-closure methods, dark speckle imaging, as well as the limitations imposed by the detectors on the performance of speckle imaging. A brief account is given of the technological innovation of adaptive-optics (AO) to compensate such atmospheric effects on the image in real time. A major advancement involves the transition from single-aperture to the dilute-aperture interferometry using multiple telescopes. Therefore, the review deals with recent developments involving ground-based, and space-based optical arrays. Emphasis is placed on the problems specific to delay-lines, beam recombination, polarization, dispersion, fringe-tracking, bootstrapping, coherencing and cophasing, and recovery of the visibility functions. The role of AO in enhancing visibilities is also discussed. The applications of interferometry, such as imaging, astrometry, and nulling are described. The mathematical intricacies of the various `post-detection' image-processing techniques are examined critically. The review concludes with a discussion of the astrophysical importance and the perspectives of interferometry.Comment: 65 pages LaTeX file including 23 figures. Reviews of Modern Physics, 2002, to appear in April issu

The Astropy Problem

The Astropy Project (http://astropy.org) is, in its own words, "a community effort to develop a single core package for Astronomy in Python and foster interoperability between Python astronomy packages." For five years this project has been managed, written, and operated as a grassroots, self-organized, almost entirely volunteer effort while the software is used by the majority of the astronomical community. Despite this, the project has always been and remains to this day effectively unfunded. Further, contributors receive little or no formal recognition for creating and supporting what is now critical software. This paper explores the problem in detail, outlines possible solutions to correct this, and presents a few suggestions on how to address the sustainability of general purpose astronomical software

Pass a Law, Any Law, Fast! State Legislative Responses to the Kelo Backlash

The Supreme Court in Kelo v. City of New London left protection of property against takings for economic development to the states. Since Kelo, thirty-seven states have enacted legislation to update their eminent domain laws. This paper is the first to theoretically and empirically analyze the factors that influence whether, in what manner, and how quickly states change their laws through new legislation. Fourteen of the thirty-seven new laws offer only weak protections against development takings. The legislative response to Kelo was responsive to measures of the backlash but only in the binary decision whether to pass any new law. The decision to enact a meaningful restriction was more a function of relevant political economy measures. States with more economic freedom, greater value of new housing construction, and less racial and income inequality are more likely to have enacted stronger restrictions, and sooner. Of the thirteen states that have not updated, Arkansas, Oklahoma and Mississippi are highly likely to do so in the future. Hawaii, Massachusetts and New York are unlikely to update ever if at all

Enthalpy of solvation correlations for organic solutes and gases dissolved in 1-propanol and tetrahydrofuran

This article discusses the enthalpy of solvation correlations for organic solutes and gases dissolved in 1-propanol and tetrahydrofuran