Crystal structure of activated CheY. Comparison with other activated receiver domains

The crystal structure of BeF(3)(-)-activated CheY, with manganese in the magnesium binding site, was determined at 2.4-A resolution. BeF(3)(-) bonds to Asp(57), the normal site of phosphorylation, forming a hydrogen bond and salt bridge with Thr(87) and Lys(109), respectively. The six coordination sites for manganese are satisfied by a fluorine of BeF(3)(-), the side chain oxygens of Asp(13) and Asp(57), the carbonyl oxygen of Asn(59), and two water molecules. All of the active site interactions seen for BeF(3)(-)-CheY are also observed in P-Spo0A(r). Thus, BeF(3)(-) activates CheY as well as other receiver domains by mimicking both the tetrahedral geometry and electrostatic potential of a phosphoryl group. The aromatic ring of Tyr(106) is found buried within a hydrophobic pocket formed by beta-strand beta4 and helix H4. The tyrosine side chain is stabilized in this conformation by a hydrogen bond between the hydroxyl group and the backbone carbonyl oxygen of Glu(89). This hydrogen bond appears to stabilize the active conformation of the beta4/H4 loop. Comparison of the backbone coordinates for the active and inactive states of CheY reveals that only modest changes occur upon activation, except in the loops, with the largest changes occurring in the beta4/H4 loop. This region is known to be conformationally flexible in inactive CheY and is part of the surface used by activated CheY for binding its target, FliM. The pattern of activation-induced backbone coordinate changes is similar to that seen in FixJ(r). A common feature in the active sites of BeF(3)(-)-CheY, P-Spo0A(r), P-FixJ(r), and phosphono-CheY is a salt bridge between Lys(109) Nzeta and the phosphate or its equivalent, beryllofluoride. This suggests that, in addition to the concerted movements of Thr(87) and Tyr(106) (Thr-Tyr coupling), formation of the Lys(109)-PO(3)(-) salt bridge is directly involved in the activation of receiver domains generally.     

Adsorption of chromium from aqueous solution by activated alumina and activated charcoal

Cr(VI) is considered to be potentially carcinogenic to humans. Removal of Cr(VI) ions from aqueous solution under different conditions was investigated using activated alumina (AA) and activated charcoal (AC) as adsorbents. Batch mode experiments were conducted to study the effects of adsorbent dose, contact time, pH, temperature and initial concentration of Cr(VI). Results showed that the adsorption of Cr(VI) depended significantly on pH and temperature. Equilibrium studies showed that Cr(VI) had a high affinity for AA at pH 4 and AC at pH 2. For AA, maximum adsorption was found at 25 degrees C, indicating exothermic adsorption, while for AC, maximum adsorption was at 40 degrees C. Freundlich and Langmuir adsorption isotherms were also applied and they showed good fits to the experimental data. The results suggest that both AA and AC could be used as effective adsorbents for the removal of Cr(VI) ions.     

Lymphokine activated killer cells

Various subpopulations of human leukocytes may be induced by lymphokines to exert cytotoxic activity. In man major histocompatibility complex non-restricted tumor cell lysis by interleukin-2 (IL-2) induced peripheral blood lymphocytes is attributed mainly to natural killer cells. These T cell receptor negative large granular lymphocytes are called lymphokine activated killer (LAK) cells. In order to explore the potential of LAK cells in tumor therapy, several clinical studies have been conducted, using IL-2 alone or in combination with ex vivo IL-2-activated peripheral blood lymphocytes. Objective responses have reproducibly been achieved only in renal cell carcinoma and malignant melanoma and were associated with considerable toxicity. In view of restricted efficacy and increasing doubts as to whether LAK cells indeed account for the in vivo observed responses, more recent strategies focus on tumor antigen specific cytotoxic T cells or tumor infiltrating lymphocytes. Successful translation of this approach into clinical practice, however, may be dependent on some basic problems of tumor immunology to be solved which were thought to be by-passed by the LAK cell approach.     

Raman activated cell sorting

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Luminescence of activated phagocytes

By means of microscope-cytofluorometry "Luman-13" intensive proper luminescence of neutrophilic granulocytes and monocytes in blood and pus, that reduce nitroblue tetrazolium has been investigated. The luminescence is localized in the cytoplasmic areas free from formazan; formazan itself does not demonstrate any luminescence. Two types of luminescence have been revealed: dark blue-blue and yellow-green. According to the luminescence spectral analysis stimulation with radiation at the wave length 360 nm produces luminescence of the I type with its maximum in the area of 450-470 nm and 520-535 nm. When stimulation with light at the wave length is 460 nm, the luminescence spectrum++ of the II type possesses the main maximum at 535 nm. Luminescence spectra of neutrophilic granulocytes and monocytes are similar. The luminescence stripe in the area 450-470 nm results from reduced pyridinenucleotides, luminescence with the wave length 520-535 nm--from oxidized flavoproteins.     

Enhanced biodegradation ofo-cresol by activated sludge in the presence of powdered activated carbon

Summary Addition of powdered activated carbon to a continuous-flow activated-sludge culture in a gas-lift bioreactor growing on o-cresol in a mineral-salts medium resulted in a reduction of the effluent o-cresol concentration during steady-state conditions as well as during shock-load conditions. During the latter conditions the activated carbon acted as a buffer. Bioregeneration of loaded activated carbon did occur, resulting in enhanced biodegradation of o-cresol. Adsorption of o-cresol by activated carbon was not sufficient to explain the total difference in effluent o-cresol concentration between the system with activated carbon and the system without activated carbon. Offprint requests to: R. J. de Jonge     

Adsorption of Cr(VI) on activated rice husk carbon and activated alumina

The possible use of activated rice husk and activated alumina as the adsorbents of Cr(VI) from synthetic solutions and the effect of operating parameters were investigated. The activated rice husk carbon was prepared thermally in two sizes 0.3 and 1.0 mm. The maximum removal of Cr(VI) occurred at pH 2 by activated rice husk and at pH 4 by activated alumina. The amounts of Cr(VI) adsorbed increased with increase in dose of both adsorbents and their contact time. The Freundlich isotherm was applied.     

Enhancement of the activated sludge process by activated carbon produced from surplus biological sludge

Surplus biological sludge from wastewater treatment operations was converted into activated carbon and then added to the aerated vessel of an activated sludge process treating phenol and glucose. The addition of activated carbon, either sludge-based or commercial, enhanced phenol removal from 58 to 98.7% and from 87 to 93% for COD with feed concentrations of 100 mg phenol l^–1 and 2500 mg COD l^–1. No differences were found between the activated sludge-activated carbon bench scale continuous reactors operating with either commercial or sludge-based activated carbon in spite of the higher adsorption capacity of the former.     

Electron-transfer activated metal-based anticancer drugs

Platinum(II)-based anticancer drugs play an essential role in the clinic today, and a number of coordination compounds with other metals are in current development as promising antitumor drugs. Probably the most prominent non-platinum metal-based drugs are those of ruthenium. Various strategies have been applied for the design of novel drugs with an improved toxicological profile, and one of them involves the preparation of metal complexes in inert high oxidation states [e.g. Pt(IV), Ru(III)]. Three platinum(IV) and two ruthenium(III) drugs have already reached clinical trials. Ideally, hypoxia-selective drugs are delivered to the target environment without prior reduction or major transformation via substitution reactions at the metal center. A (selective) reduction has been proposed to activate the prodrugs by formation of active species, which react with the target more readily and lead ultimately to apoptosis. Investigations on the electrochemical behavior of platinum(IV) and ruthenium(III) cytotoxins and the establishment of preliminary structure-property relationships are therefore of current importance. Herein, we present recent results in the field of metal-centered electron-transfer activated Ru(III), Pt(IV) and Co(III) drugs with regard to design and targeting strategies, prediction of redox potentials in aqueous medium, labilization and enhanced reactivity with potential biological targets upon reduction, and correlations between electrochemical parameters and anticancer activity.     

Aeration of concentrated activated sludge

A multiphase project has been planned to develop a new biological process capable of economically treating high BOD wastes. Herein is presented the results of the first phase of the program, in which the feasibility of growing concentrated microbial cultures was investigated and the oxygen and power requirements for maintaining such cultures were determined. An example is given of the scale-up of power requirements for oxygen transfer in a prototype system.     

Structure of activated sludge floes

Relatively large activated sludge floes (larger than about 100 mum) were stabilized, using a histological tissue specimen preparation procedure, and then were sliced into sections of 3 to 6 mum thick. The study of these sections, after staining, revealed the internal structure of the activated sludge floes. No uniformity of this structure was found. The distribution of microorganisms and of extracellular polymers (EPs) in the floes varied randomly on the plane of the sections and along the dimension perpendicular to the plane, leaving large water channels and reservoirs in some of the floes. The lack of a characteristic size for the water gaps in the floes and a general self-similar appearance of the sections suggested that the activated sludge floes might be characterized by the fractal concept within a certain size limit. Direct observation of the interior of the floes indicated an abundant presence of extracellular polymers (EPs) in amorphous forms, surrounding microorganisms in most of the floes.     

Enzymatic activities of activated sludge

Summary As a potential measure of active biomass in an automated and continuous system, enzyme activities of activated sludge were sought that would allow a rapid and accurate determination. Using p-nitrophenylphosphate, bis-p-nitrophenylphosphate, p-nitrophenyl-- and -D-gluco- or galactopyranosides, and the p-nitroanilides of L-alanine, L-leucine, L-lysine, L-glutamic acid and L-phenylalanine, we could establish that the corresponding phosphatase, cyclic phosphodiesterase, glycosidase and aminopeptidase activities can be conveniently tested with diluted activated sludge within 10–20 min incubation at 30°C. The reactions were linear with time and concentration of activated sludge. The specific activities were in the range from 1 to 50 nmol substrate cleaved/min/mg protein at 30°C. They were diminished by starvation or poisoning with zinc powder, ZnCl₂, HgCl₂ or KCN. The enzymes were over 95% sedimented together with the flocs. Comparable activities were found in sludges from three independent sewage works in the Munich area. At the same time, dehydrogenase activities and ATP contents were investigated under several conditions.     

Spatial-temporal reorganization of activated integrins

Integrin receptors play important roles in cell adhesion and tumor metastasis. The coupling of mechanical sensing and biochemical ligation is known to collectively regulate the activation of integrin receptors. Recently, oligomerization of activated integrins has been considered as the primordial signature of cytoskeletal remodeling and the initiation of various downstream signals, such as focal and fibrillar adhesions. However, spatio-temporal reorganization of activated integrins and associated proteins remains poorly understood. Here, we summarized the recent discovery of sequential biophysical events of integrin activation during early adhesion formation. Using the cyclic Arg-Gly-Asp (RGD) peptide as a mobile ligand on supported lipid membranes, a series of previously unreported events were observed following integrin αvβ3 clustering and cell spreading, including a long-range lateral translocation of the integrin clusters. With initial clustering, localized actin polymerization occurred in a Src family kinase dependent manner. Clustering of liganded integrins recruits various adaptor proteins and serves as a reaction core for mechanobiological activities. In addition, there are future possibilities to investigate the role of other synergetic interactions with the activated integrin receptors.     

Nucleoside transport in activated macrophages

The study of [3H]-uridine uptake by mouse peritoneal macrophages showed that this is an active, temperature- and protein synthesis-dependent phenomenon, which is early altered when are exposed to a variety of stimuli. Murine recombinant interferon-gamma, a stimulus able to activate macrophage and to induce the production of tumor necrosis factor-alpha, within few hours markedly increased [3H]-uridine uptake by mouse macrophage. Other stimuli devoid of activation capacity, such as inert phagocytable latex beads, did not affect this phenomenon, which appeared to be related to macrophage activation. The increase in [3H]-uridine uptake may be an useful phenomenon in studying the early biochemical events associated with macrophage activation.