Translocation of the catalytic domain of diphtheria toxin across planar phospholipid bilayers by its own T domain. (1/307)

The T domain of diphtheria toxin is known to participate in the pH-dependent translocation of the catalytic C domain of the toxin across the endosomal membrane, but how it does so, and whether cellular proteins are also required for this process, remain unknown. Here, we report results showing that the T domain alone is capable of translocating the entire C domain across model, planar phospholipid bilayers in the absence of other proteins. The T domain therefore contains the entire molecular machinery for mediating transfer of the catalytic domain of diphtheria toxin across membranes.  (+info)

Field trials of the rodenticide gophacide against wild house mice (Mus musculus L.). (2/307)

The acute rodenticide gophacide was tested against urban infestations of the house mouse (Mus musculus L.) and treatment success was assessed from the results of census baitings conducted before and after each treatment. Seven of eight populations of mice living in premises where alternative food supplies were limited were successfully controlled when medium oatmeal bait containing gophacide at 0.1% was laid directly for 4 days. In further treatments against mice inhabiting more complex environments and having greater access to other foods, the performance of gophacide at 0.1% and at 0.25% in a wholemeal flour/pinhead oatmeal/corn oil bait was compared with that of zinc phosphide at 3.0% in the same bait-base. The poison treatments were conducted for 1 or 4 days and always after 3 days pre-baiting. Treatment success varied considerably irrespective of the type of treatment or of the poison used. In general, however, gophacide proved to be as effective as zinc phosphide for the control of mice.  (+info)

Disulfide structure of the pheromone binding protein from the silkworm moth, Bombyx mori. (3/307)

Disulfide bond formation is the only known posttranslational modification of insect pheromone binding proteins (PBPs). In the PBPs from moths (Lepidoptera), six cysteine residues are highly conserved at positions 19, 50, 54, 97, 108 and 117, but to date nothing is known about their respective linkage or redox status. We used a multiple approach of enzymatic digestion, chemical cleavage, partial reduction with Tris-(2-carboxyethyl)phosphine, followed by digestion with endoproteinase Lys-C to determine the disulfide connectivity in the PBP from Bombyx mori (BmPBP). Identification of the reaction products by on-line liquid chromatography-electrospray ionization mass spectrometry (LC/ESI-MS) and protein sequencing supported the assignment of disulfide bridges at Cys-19-Cys-54, Cys-50-Cys-108 and Cys-97-Cys-117. The disulfide linkages were identical in the protein obtained by periplasmic expression in Escherichia coli and in the native BmPBP.  (+info)

A comparative field trial, conducted without pre-treatment census baiting, of the rodenticides zinc phosphide, thallium sulphate and gophacide against Rattus norvegicus. (4/307)

The effectiveness of the single-dose poison treatments of farm rat infestations, analysed by comparing the weights of the post-treatment census bait takes in covariance with the weights of the prebait takes, showed that treatments with 2-5% zinc phosphide, 0-3% thallium sulphate or 0-3% gophacide were equally effective and significantly better than were treatments with 1% zinc phosphide or 0-1% thallium sulphate. The methodology and sensitivity of different analyses are also considered.  (+info)

Novel role for the NMDA receptor redox modulatory site in the pathophysiology of seizures. (5/307)

Redox-active compounds modulate NMDA receptors (NMDARs) such that reduction of NMDAR redox sites increases, and oxidation decreases, NMDAR-mediated activity. Because NMDARs contribute to the pathophysiology of seizures, redox-active compounds also may modulate seizure activity. We report that the oxidant 5, 5'-dithio-bis(2-nitrobenzoic acid) (DTNB) and the redox cofactor pyrroloquinoline quinone (PQQ) suppressed low Mg(2+)-induced hippocampal epileptiform activity in vitro. Additionally, in slices exposed to 4-7 microM bicuculline, DTNB and PQQ reversed the potentiation of evoked epileptiform responses by the reductants dithiothreitol and Tris(2-carboxyethyl)phosphine (TCEP). NMDA-evoked whole-cell currents in CA1 neurons in slices were increased by TCEP and subsequently decreased by DTNB or PQQ at the same concentrations that modulated epileptiform activity. However, DTNB and PQQ had little effect on baseline NMDA-evoked currents in control medium, and PQQ did not alter NMDAR-dependent long-term potentiation. In contrast, in slices returned to control medium after low Mg(2+)-induced ictal activity, DTNB significantly inhibited NMDAR-mediated currents, indicating endogenous reduction of NMDAR redox sites under this epileptogenic condition. These data suggested that PQQ and DTNB suppressed spontaneous ictal activity by reversing pathological NMDAR redox potentiation without inhibiting physiological NMDAR function. In vivo, PQQ decreased the duration of chemoconvulsant-induced seizures in rat pups with no effect on baseline behavior. Our results reveal endogenous potentiation of NMDAR function via mass reduction of redox sites as a novel mechanism that may enhance epileptogenesis and facilitate the transition to status epilepticus. The results further suggest that redox-active compounds may have therapeutic use by reversing NMDAR-mediated pathophysiology without blocking physiological NMDAR function.  (+info)

Cell surface engineering by a modified Staudinger reaction. (6/307)

Selective chemical reactions enacted within a cellular environment can be powerful tools for elucidating biological processes or engineering novel interactions. A chemical transformation that permits the selective formation of covalent adducts among richly functionalized biopolymers within a cellular context is presented. A ligation modeled after the Staudinger reaction forms an amide bond by coupling of an azide and a specifically engineered triarylphosphine. Both reactive partners are abiotic and chemically orthogonal to native cellular components. Azides installed within cell surface glycoconjugates by metabolism of a synthetic azidosugar were reacted with a biotinylated triarylphosphine to produce stable cell-surface adducts. The tremendous selectivity of the transformation should permit its execution within a cell's interior, offering new possibilities for probing intracellular interactions.  (+info)

Fatal aluminum phosphide poisoning. (7/307)

A 39-year-old man committed suicide by ingestion of aluminum phosphide, a potent mole pesticide, which was available at the victim's workplace. The judicial authority ordered an autopsy, which ruled out any other cause of death. The victim was discovered 10 days after the ingestion of the pesticide. When aluminum phosphide comes into contact with humidity, it releases large quantities of hydrogen phosphine (PH3), a very toxic gas. Macroscopic examination during the autopsy revealed a very important asphyxia syndrome with major visceral congestion. Blood, urine, liver, kidney, adrenal, and heart samples were analyzed. Phosphine gas was absent in the blood and urine but present in the brain (94 mL/g), the liver (24 mL/g), and the kidneys (41 mL/g). High levels of phosphorus were found in the blood (76.3 mg/L) and liver (8.22 mg/g). Aluminum concentrations were very high in the blood (1.54 mg/L), brain (36 microg/g), and liver (75 microg/g) compared to the usual published values. Microscopic examination revealed congestion of all the organs studied and obvious asphyxia lesions in the pulmonary parenchyma. All these results confirmed a diagnosis of poisoning by aluminum phosphide. This report points out that this type of poisoning is rare and that hydrogen phosphine is very toxic. The phosphorus and aluminum concentrations observed and their distribution in the different viscera are discussed in relation to data in the literature.  (+info)

Interaction between the lipoamide-containing H-protein and the lipoamide dehydrogenase (L-protein) of the glycine decarboxylase multienzyme system. 1. Biochemical studies. (8/307)

Lipoamide dehydrogenase or dihydrolipoamide dehydrogenase (EC 1.8.1. 4) is the E3-protein component of the mitochondrial 2-oxoacid dehydrogenase multienzyme complexes. It is also the L-protein component of the glycine decarboxylase system. Although the enzymology of this enzyme has been studied exhaustively using free lipoamide as substrate, no data are available concerning the kinetic parameters of this enzyme with its physiological substrates, the dihydrolipoyl domain of the E2 component (dihydrolipoyl acyltransferase) of the 2-oxoacid dehydrogenase multienzyme complexes or the dihydrolipoyl H-protein of the mitochondrial glycine decarboxylase. In this paper, we demonstrate that Tris(2-carboxyethyl)phosphine, a specific disulfide reducing agent, allows a continuous reduction of the lipoyl group associated with the H-protein during the course of the reaction catalysed by the L-protein. This provided a valuable new tool with which to study the catalytic properties of the lipoamide dehydrogenase. The L-protein displayed a much higher affinity for the dihydrolipoyl H-protein than for free dihydrolipoamide. The oxidation of the dihydrolipoyl H-protein was not affected by the presence of structurally related analogues (apoH-protein or octanoylated H-protein). In marked contrast, these analogues strongly and competitively inhibited the decarboxylation of the glycine molecule catalysed by the P-protein component of the glycine decarboxylase system. Small unfolded proteolytic fragments of the H-protein, containing the lipoamide moiety, displayed Km values for the L-protein close to that found for the H-protein. On the other hand, these fragments were not able to promote the decarboxylation of the glycine in the presence of the P-protein. New highly hydrophilic lipoate analogues were synthesized. All of them showed Km and kcat/Km values very close to that found for the H-protein. From our results we concluded that no structural interaction is required for the L-protein to catalyse the oxidation of the dihydrolipoyl H-protein. We discuss the possibility that one function of the H-protein is to maintain a high concentration of the hydrophobic lipoate molecules in a nonmicellar state which would be accessible to the catalytic site of the lipoamide dehydrogenase.  (+info)