Conserved Aspartate Residues and Phosphorylation in Signal Transduction by the Chemotaxis Protein CheY

The CheY protein is phosphorylated by CheA and dephosphorylated by CheZ as part of the chemotactic signal transduction pathway in Escherichia coli. Phosphorylation of CheY has been proposed to occur on an aspartate residue. Each of the eight aspartate residues of CheY was replaced by using site-directed mutagenesis. Substitutions at Asp-12, Asp-13, or Asp-57 resulted in loss of chemotaxis. Most of the mutant CheY proteins were still phosphorylated by CheA but exhibited modified biochemical properties, including reduced ability to accept phosphate from CheA, altered phosphate group stability, and/or resistance to CheZ-mediated dephosphorylation. The properties of CheY proteins bearing a substitution at position 57 were most aberrant, consistent with the hypothesis that Asp-57 is the normal site of acyl phosphate formation. Evidence for an alternate site of phosphorylation in the Asp-57 mutants is presented. Phosphorylated CheY is believed to cause tumbling behavior. However, a dominant mutant CheY protein that was not phosphorylated in vitro caused tumbling in vivo in the absence of CheA. This phenotype suggests that the role of phosphorylation in the wild-type CheY protein is to stabilize a transient conformational change that can generate tumbling behavior

Optimization of payload mass placement in a dual keel space station

In order to keep a Space Station in a stable low-Earth orbit, angular momentum storage and translational attitude control systems will have to be used. In order to minimize the size of these attitude control systems, the induced gravity gradient torque effects will have to be minimized. This can be done by minimizing the cross-products of inertia of the Station through the management of payload placement with the Station geometry. A derived and automated methodology is presented which utilizes mathematical nonlinear programming techniques. An optimal arrangement of a set of five payloads on a Dual Keel Space Station was found that minimized the cross products of inertia and thus the required controllability resources

Uncoupled Phosphorylation and Activation in Bacterial Chemotaxis - The 2.3 Å structure of an aspartate to lysine mutant at position 13 of CheY

An aspartate to lysine mutation at position 13 of the chemotaxis regulatory protein CheY causes a constitutive tumbly phenotype when expressed at high copy number in vivo even though the mutant protein is not phosphorylatable. These properties suggest that the D13K mutant adopts the active, signaling conformation of CheY independent of phosphorylation, so knowledge of its structure could explain the activation mechanism of CheY. The x-ray crystallographic structure of the CheY D13K mutant has been solved and refined at 2.3 Å resolution to an R-factor of 14.3%. The mutant molecule shows no significant differences in backbone conformation when compared with the wild-type, Mg2+-free structure, but there are localized changes within the active site. The side chain of lysine 13 blocks access to the active site, whereas its epsilon -amino group has no bonding interactions with other groups in the region. Also in the active site, the bond between lysine 109 and aspartate 57 is weakened, and the solvent structure is perturbed. Although the D13K mutant has the inactive conformation in the crystalline form, rearrangements in the active site appear to weaken the overall structure of that region, potentially creating a metastable state of the molecule. If a conformational change is required for signaling by CheY D13K, then it most likely proceeds dynamically, in solution

Marketing Timber from the Private Woodland

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INITIAL CHARACTERIZATION OF MAJOR HISTOCOMPATIBILITY COMPLEX (MHC) CLASS IIB EXON 2 IN AN ENDANGERED RATTLESNAKE, THE EASTERN MASSASAUGA (SISTRURUS CATENATUS)

Genes of the major histocompatibility complex (MHC) play an important role in the vertebrate immune system and exhibit remarkably high levels of polymorphism, maintained by strong balancing selection. While the conservation implications of MHC variation have been explored in a variety of vertebrates, non-avian reptiles (most notably snakes) have received less attention. To address this gap and take the first steps toward more extensive population-level analyses, we cloned and sequenced MHC IIB exon 2 in an endangered rattlesnake, the Eastern Massasauga (Sistrurus catenatus). Based on three individuals, we found evidence of at least four putatively functional loci. These sequences exhibited relatively high levels of variation and significantly higher rates of nonsynonymous to synonymous substitutions, especially within the antigen-binding sites, indicating strong positive selection. Phylogenetic analysis revealed a pattern of trans-species polymorphism, also suggesting positive selection. These results contribute to our understanding of MHC variation in non-avian reptiles and form a basis for more studies of MHC variation in snakes of conservation concern

The 'anti-economics' of the European common market

I would like to preface my comments by extending a personal but public note of appreciation to the administrators, faculty and students of the Royal University for inviting me to visit and address this distinguished audience of scholars, business men and government officers, this evening on the theme of the 'Anti-Economics' of the European Common Market and its relation to the tax harmonization program of the Community. Perhaps the title of my talk warrants clarification? What is precisely meant by the 'anti-economics' of the European Economic Community for one; and what is meant by 'tax harmonization' for the other? And even more important, why should this distinguished gathering be interested in either one of them?peer-reviewe

Pengaruh Mekanisme Tata Kelola Perusahaan dan Struktur Kepemilikan terhadap Struktur Modal (Studi Empiris pada Perusahaan Manufaktur yang Terdaftar di Bursa Efek Indonesia pada Tahun 2011-2013)

The purpose of this study is to examine the influence of corporate governance mechanisms and ownership structure to capital structure. The dependent variable is capital structure which is proxied as debt ratio (DR). Independent variable are corporate governance mechanism which is proxied by size of the board of commissioners, size of the board of independent commissioners, size of the board of directors, size of the audit committee. Ownership structure which is proxied by managerial ownership and institutional ownership. This study was used secondary data from annual reports of manufacturing companies which were listed on Indonesia Stock Exchange in 2011-2013. Samples were 40 manufacturing companies. This study used purposive sampling method and multiple linear regression as the analysis method. Before being conducted by regression test, it was examined by using the classical assumption tests. The results of this study indicate that the size of the board of commisioners, size of the board of independent commisioners, size of board committee, and institusional ownership did not have significant influence to the agency cost. The size of the board of directors and managerial ownership have significant influence to capital structure

Pre-bit hackamore training (1993)

Hackamores are used to start colts in training. An untrained colt makes many mistakes, and the trainer needs to correct them. This publication gives instructions on how to use a hackamore

Plasma Corticotropin-Releasing Factor Receptors and B7-2⁺ Extracellular Vesicles in Blood Correlate with Irritable Bowel Syndrome Disease Severity.

Extracellular vesicles (EVs) are composed of bilayer membranes that are released by different cell types and are present in bodily fluids, such as blood, urine, and bile. EVs are thought to play a key role in intracellular communication. Based on their size and density, EVs are classified into small, medium, or large EVs. Cargo composition in EVs reflects physiological changes in health and disease. Patients with irritable bowel syndrome (IBS) exhibit visceral hypersensitivity and mood disorders. Stressful episodes often precede disease symptoms in IBS patients. Stress-induced symptoms include, but are not limited to, abdominal pain and mood swings. Perceived stress responses are mediated by two known G protein-coupled receptors (GPCRs), corticotropin-releasing factor receptor 1 and 2 (CRFRs). CRFRs belong to the Class B secretin receptor family of GPCRs. Here, we show that CRFRs were present in human and murine plasma, and in EVs purified from mouse serum. CRFRs were present in plasma from IBS patients and healthy controls. EVs secreted from immune cells influence both adaptive and innate immune responses via exchange of EVs between different immune cell types. B7-2 (CD86), a plasma membrane antigen-presenting protein, is present on EVs secreted from dendritic, B-, and mast cells, whereas CD9 is present on EVs secreted from dendritic and intestinal epithelial cells. We found that plasma CRFR levels positively correlated with B7-2+ EVs (R = 0.8597, p < 0.0001), but no association was seen with CD9+ EVs. Plasma CRFRs expression negatively correlated with IBS severity scores. Our data suggests that plasma EVs from immune cells carry CRFRs as cargos and influence cell-cell communication in health and disease

Retinal degeneration is rescued in transgenic rd mice by expression of the cGMP phosphodiesterase ß subunit

The ß subunit of the cGMP phosphodiesterase (PDE) gene has been identified as the candidate gene for retinal degeneration in the rd mouse. To study the molecular mechanisms underlying degeneration and the potential for gene repair, we have expressed a functional bovine cGMP PDE ß subunit in transgenic rd mice. One transgenic mouse line showed complete photoreceptor rescue across the entire span of the retina. A second independently derived line showed partial rescue in which photoreceptors in the superior but not the inferior hemisphere of the retina were rescued. In the latter animals, intermediate stages of degeneration were observed in the transition zone between rescued and diseased photoreceptors. Pathologic changes in the retina ranged from vesiculation of the basalmost outer segment discs in otherwise structurally intact rod cells to photoreceptors with highly disorganized outer segments and intact inner segments. Totally or partially rescued retinas showed a corresponding restoration of cGMP PDE activity, whereas nonrescued retinas had minimal enzyme activity, characteristic of the rd phenotype. These transgenic animals provide models for studying the molecular basis of retinal degenerative disease and conclusively demonstrate that the phenotype of rd mice is produced by a defect in the ß subunit of cGMP PDE