The backbone structure of the thermophilic Thermoanaerobacter tengcongensis ribose binding protein is essentially identical to its mesophilic E. coli homolog

<p>Abstract</p> <p>Background</p> <p>Comparison of experimentally determined mesophilic and thermophilic homologous protein structures is an important tool for understanding the mechanisms that contribute to thermal stability. Of particular interest are pairs of homologous structures that are structurally very similar, but differ significantly in thermal stability.</p> <p>Results</p> <p>We report the X-ray crystal structure of a <it>Thermoanaerobacter tengcongensis </it>ribose binding protein (tteRBP) determined to 1.9 Å resolution. We find that tteRBP is significantly more stable (^<it>app</it><it>T</it>_{<it>m </it>}value ~102°C) than the mesophilic <it>Escherichia coli </it>ribose binding protein (ecRBP) (^<it>app</it><it>T</it>_{<it>m </it>}value ~56°C). The tteRBP has essentially the identical backbone conformation (0.41 Å RMSD of 235/271 C_αpositions and 0.65 Å RMSD of 270/271 C_αpositions) as ecRBP. Classification of the amino acid substitutions as a function of structure therefore allows the identification of amino acids which potentially contribute to the observed thermal stability of tteRBP in the absence of large structural heterogeneities.</p> <p>Conclusion</p> <p>The near identity of backbone structures of this pair of proteins entails that the significant differences in their thermal stabilities are encoded exclusively by the identity of the amino acid side-chains. Furthermore, the degree of sequence divergence is strongly correlated with structure; with a high degree of conservation in the core progressing to increased diversity in the boundary and surface regions. Different factors that may possibly contribute to thermal stability appear to be differentially encoded in each of these regions of the protein. The tteRBP/ecRBP pair therefore offers an opportunity to dissect contributions to thermal stability by side-chains alone in the absence of large structural differences.</p

Could This Train Make It Through: The Law and Strategy of the Gold Train Case

In 1944-45, the Nazis seized personal belongings of the Hungarian Jewish population and dispatched some of the most valuable of them on a train. The United States Army took control of this Gold Train and gave reassurances that it would keep the valuables safe. However, the items were plundered by individual soldiers, including officers, and diverted to various uses. After decades of dormancy, a Presidential Commission exposed the facts, but the government still did not right the wrong — until there was litigation. The Gold Train case (Rosner v. United States) represents a measure of justice for the victimized community of Hungarian Jewish Holocaust survivors. This case is one of the most successful human rights class actions ever brought against the United States. It teaches important lessons regarding future human rights cases, especially those against the United States. These lessons concern both the legal doctrines in such cases and strategic questions about how to mobilize the public\u27s sympathy for human rights victims injured by the United States abroad. Survivors and descendants of Hungarian Jews brought this class action against the United States to seek restitution because their valuables were seized by the Nazis in 1945 and subsequently loaded on a train which came into United States Army control. The valuables were disposed of by grants to international organizations, sale, waste, and looting, rather than being returned to the owners. Strategically, Hungarian Jewry, as Holocaust-era victims, had the sympathy of American Jewry, a group whose voice would help bring about a just settlement. The Rosner settlement obliged the United States to devote $25 million to the needs of this class of Holocaust victims. There is much more to the Gold Train case than meets the eye, and the litigation took many unanticipated turns over five years of preparation, research, litigation, negotiation, and settlement. Doctrinally and strategically the route to obtaining justice was anything but simple. The underlying events occurred in far flung locales — Hungary, Austria, France, London, New York, Budapest, Israel, and Washington, DC, among others over 55 years before the case was brought. Both factual and legal questions posed major challenges

Could This Train Make It Through: The Law and Strategy of the Gold Train Case

In 1944-45, the Nazis seized personal belongings of the Hungarian Jewish population and dispatched some of the most valuable of them on a train. The United States Army took control of this Gold Train and gave reassurances that it would keep the valuables safe. However, the items were plundered by individual soldiers, including officers, and diverted to various uses. After decades of dormancy, a Presidential Commission exposed the facts, but the government still did not right the wrong — until there was litigation. The Gold Train case (Rosner v. United States) represents a measure of justice for the victimized community of Hungarian Jewish Holocaust survivors. This case is one of the most successful human rights class actions ever brought against the United States. It teaches important lessons regarding future human rights cases, especially those against the United States. These lessons concern both the legal doctrines in such cases and strategic questions about how to mobilize the public\u27s sympathy for human rights victims injured by the United States abroad. Survivors and descendants of Hungarian Jews brought this class action against the United States to seek restitution because their valuables were seized by the Nazis in 1945 and subsequently loaded on a train which came into United States Army control. The valuables were disposed of by grants to international organizations, sale, waste, and looting, rather than being returned to the owners. Strategically, Hungarian Jewry, as Holocaust-era victims, had the sympathy of American Jewry, a group whose voice would help bring about a just settlement. The Rosner settlement obliged the United States to devote $25 million to the needs of this class of Holocaust victims. There is much more to the Gold Train case than meets the eye, and the litigation took many unanticipated turns over five years of preparation, research, litigation, negotiation, and settlement. Doctrinally and strategically the route to obtaining justice was anything but simple. The underlying events occurred in far flung locales — Hungary, Austria, France, London, New York, Budapest, Israel, and Washington, DC, among others over 55 years before the case was brought. Both factual and legal questions posed major challenges

Ligand-induced conformational changes in a thermophilic ribose-binding protein

<p>Abstract</p> <p>Background</p> <p>Members of the periplasmic binding protein (PBP) superfamily are involved in transport and signaling processes in both prokaryotes and eukaryotes. Biological responses are typically mediated by ligand-induced conformational changes in which the binding event is coupled to a hinge-bending motion that brings together two domains in a closed form. In all PBP-mediated biological processes, downstream partners recognize the closed form of the protein. This motion has also been exploited in protein engineering experiments to construct biosensors that transduce ligand binding to a variety of physical signals. Understanding the mechanistic details of PBP conformational changes, both global (hinge bending, twisting, shear movements) and local (rotamer changes, backbone motion), therefore is not only important for understanding their biological function but also for protein engineering experiments.</p> <p>Results</p> <p>Here we present biochemical characterization and crystal structure determination of the periplasmic ribose-binding protein (RBP) from the hyperthermophile <it>Thermotoga maritima </it>in its ribose-bound and unliganded state. The <it>T. maritima </it>RBP (tmRBP) has 39% sequence identity and is considerably more resistant to thermal denaturation (^<it>app</it><it>T</it>_{<it>m </it>}value is 108°C) than the mesophilic <it>Escherichia coli </it>homolog (ecRBP) (^<it>app</it><it>T</it>_{<it>m </it>}value is 56°C). Polar ligand interactions and ligand-induced global conformational changes are conserved among ecRBP and tmRBP; however local structural rearrangements involving side-chain motions in the ligand-binding site are not conserved.</p> <p>Conclusion</p> <p>Although the large-scale ligand-induced changes are mediated through similar regions, and are produced by similar backbone movements in tmRBP and ecRBP, the small-scale ligand-induced structural rearrangements differentiate the mesophile and thermophile. This suggests there are mechanistic differences in the manner by which these two proteins bind their ligands and are an example of how two structurally similar proteins utilize different mechanisms to form a ligand-bound state.</p

Radio frequency plasma processing effects on the emission characteristics of a MeV electron beam cathode

Experiments have proven that surface contaminants on the cathode of an electron beam diode influence electron emission current and impedance collapse. This letter reports on an investigation to reduce parasitic cathode current loss and to increase high voltage hold off capabilities by reactive sputter cleaning of contaminants. Experiments have characterized effective radio frequency (rf) plasma processing protocols for high voltage anode–cathode (A–K) gaps using a two-stage argon/oxygen and argon rf plasma discharge. Time-resolved optical emission spectroscopy measures contaminant (hydrogen) and bulk cathode (aluminum) plasma emission versus transported axial electron beam current turn on. Experiments were performed at accelerator parameters: V = −0.7V=−0.7 to −1.1 MV,−1.1MV, I(diode)=3–30 kA,I(diode)=3–30kA, and pulse length=0.4–1.0 μs.length=0.4–1.0μs. Experiments using a two-stage low power (100 W) argon/oxygen rf discharge followed by a higher power (200 W) pure argon rf discharge yielded an increase in cathode turn-on voltage required for axial current emission from 662±174 kV662±174kV to 981±97 kV.981±97kV. The turn-on time of axial current was increased from 100±22100±22 to 175±42 ns.175±42ns. © 1999 American Institute of Physics.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/69353/2/APPLAB-75-1-31-1.pd

Individual quality assessment of autografting by probability estimation for clinical endpoints: a prospective validation study from the European group for blood and marrow transplantation.

The aim of supportive autografting is to reduce the side effects from stem cell transplantation and avoid procedure-related health disadvantages for patients at the lowest possible cost and resource expenditure. Economic evaluation of health care is becoming increasingly important. We report clinical and laboratory data collected from 397 consecutive adult patients (173 non-Hodgkin lymphoma, 30 Hodgkin lymphoma, 160 multiple myeloma, 7 autoimmune diseases, and 28 acute leukemia) who underwent their first autologous peripheral blood stem cell transplantation (PBSCT). We considered primary endpoints evaluating health economic efficacy (eg, antibiotic administration, transfusion of blood components, and time in hospital), secondary endpoints evaluating toxicity (in accordance with Common Toxicity Criteria), and tertiary endpoints evaluating safety (ie, the risk of regimen-related death or disease progression within the first year after PBSCT). A time-dependent grading of efficacy is proposed with day 21 for multiple myeloma and day 25 for the other disease categories (depending on the length of the conditioning regimen) as the acceptable maximum time in hospital, which together with antibiotics, antifungal, or transfusion therapy delineates four groups: favorable (≤7 days on antibiotics and no transfusions; ≤21 [25] days in hospital), intermediate (from 7 to 10 days on antibiotics and 7 days on antibiotics, >3 but 30/34 days in hospital after transplantation), and very unfavorable (>10 days on antibiotics, >6 transfusions; >30 to 34 days in hospital). The multivariate analysis showed that (1) PBSC harvests of ≥4 × 106/kg CD34 + cells in 1 apheresis procedure were associated with a favorable outcome in all patient categories except acute myelogenous leukemia and acute lymphoblastic leukemia (P = .001), (2) ≥5 × 106/kg CD34 + cells infused predicted better transplantation outcome in all patient categories (P 500 mL) (P = .002), and (5) patients with a central venous catheter during both collection and infusion of PBSC had a more favorable outcome post-PBSCT than peripheral access (P = .007). The type of mobilization regimen did not affect the outcome of auto-PBSCT. The present study identified predictive variables, which may be useful in future individual pretransplantation probability evaluations with the goal to improve supportive care

Imaging Primary Mouse Sarcomas After Radiation Therapy Using Cathepsin-Activatable Fluorescent Imaging Agents

Purpose: Cathepsin-activated fluorescent probes can detect tumors in mice and in canine patients. We previously showed that these probes can detect microscopic residual sarcoma in the tumor bed of mice during gross total resection. Many patients with soft tissue sarcoma (STS) and other tumors undergo radiation therapy (RT) before surgery. This study assesses the effect of RT on the ability of cathepsin-activated probes to differentiate between normal and cancerous tissue. Methods and Materials: A genetically engineered mouse model of STS was used to generate primary hind limb sarcomas that were treated with hypofractionated RT. Mice were injected intravenously with cathepsin-activated fluorescent probes, and various tissues, including the tumor, were imaged using a hand-held imaging device. Resected tumor and normal muscle samples were harvested to assess cathepsin expression by Western blot. Uptake of activated probe was analyzed by flow cytometry and confocal microscopy. Parallel in vitro studies using mouse sarcoma cells were performed. Results: RT of primary STS in mice and mouse sarcoma cell lines caused no change in probe activation or cathepsin protease expression. Increasing radiation dose resulted in an upward trend in probe activation. Flow cytometry and immunofluorescence showed that a substantial proportion of probe-labeled cells were CD11b-positive tumor-associated immune cells. Conclusions: In this primary murine model of STS, RT did not affect the ability of cathepsin-activated probes to differentiate between tumor and normal muscle. Cathepsin-activated probes labeled tumor cells and tumor-associated macrophages. Our results suggest that it would be feasible to include patients who have received preoperative RT in clinical studies evaluating cathepsin-activated imaging probes.Damon Runyon Cancer Research Foundation (Damon Runyon-Rachleff Innovation Award

Seed Cone Anatomy of Cheirolepidiaceae (Coniferales): Reinterpreting Pararaucaria patagonica Wieland

Premise of the study: Seed cone morphology and anatomy reflect some of the most important changes in the phylogeny and evolutionary biology of conifers. Reexamination of the enigmatic Jurassic seed cone Pararaucaria patagonica reveals previously unknown systematically informative characters that demonstrate affinities with the Cheirolepidiaceae. This paper documents, for the first time, internal anatomy for seed cones of this important extinct Mesozoic conifer family, which may represent the ghost lineage leading to modern Pinaceae. Methods: Morphology and anatomy of cones from the Jurassic La Matilde Formation in Patagonia are described from a combination of polished wafers and thin section preparations. New photographic techniques are employed to reveal histological details of thin sections in which organic cell wall remains are not preserved. Specific terminology for conifer seed cones is proposed to help clarify hypotheses of homology for the various structures of the cones. Key results: Specimens are demonstrated to have trilobed ovuliferous scale tips along with a seed enclosing pocket of ovuliferous scale tissue. Originally thought to represent a seed wing in P. patagonica, this pocket-forming tissue is comparable to the flap of tissue covering seeds of compressed cheirolepidiaceous cones and is probably the most diagnostic character for seed cones of the family. Conclusions: Pararaucaria patagonica is assigned to Cheirolepidiaceae, documenting anatomical features for seed cones of the family and providing evidence for the antiquity of pinoid conifers leading to the origin of Pinaceae. A list of key morphological and anatomical characters for seed cones of Cheirolepidiaceae is developed to facilitate assignment of a much broader range of fossil remains to the family. This confirms the presence of Cheirolepidiaceae in the Jurassic of the Southern Hemisphere, which was previously suspected from palynological records.Keywords: Conifers, Seed cones, Cheirolepidiaceae, Jurassic, Pararaucaria, Patagoni