Kinetic studies of a soluble alpha beta complex of nitrate reductase A from Escherichia coli. Use of various alpha beta mutants with altered beta subunits

A soluble alpha beta complex of nitrate reductase can be obtained from a strain of Escherichia coli that lacks the narI gene and expresses only the alpha and beta subunits. The beta subunit contains four Fe-S centres and the alpha subunit contains the molybdenum cofactor, which is the site at which nitrate is reduced. Despite the lack of the gamma subunit of the complete enzyme, this complex can still catalyse the reduction of nitrate with artificial electron donors such as benzyl viologen, so that it is suitable for studying the transfer of electrons between these two types of redox centre. To examine whether the electrons from reduced benzyl viologen are initially delivered to the Fe-S centres, or directly to the molybdenum cofactor, or both, we have studied the steady-state kinetics and the binding of benzyl viologen to the alpha beta complex and mutants alpha beta* with altered beta subunits. Reduction of the enzyme by reduced benzyl viologen in the absence of nitrate showed that all four Fe-S centres and the molybdenum cofactor could be reduced. Two classes of site with different equilibrium constants could be distinguished. The kinetic results suggest that benzyl viologen supplies its electrons directly to the molybdenum cofactor, at a rate showing a hyperbolic dependence on the square of the concentration of the electron donor. A reaction mechanism is proposed for the reduction of nitrate catalysed by the alpha beta complex of nitrate reductase with artificial electron donors.     

Influence of Manufacturing and Ripening Conditions on the Survival of Enterobacteriaceae in Manchego Cheese

The effects of cooking temperature, time and temperature of brine-salting, and temperature of ripening on the behavior of Enterobacteriaceae in Manchego cheese were studied. Fifty lots of cheese from raw ewe's milk manufactured and ripened under different conditions were investigated throughout a 60-day ripening period. Differences in pH values due to temperature-dependent whey retention accounted for the effect of cooking temperature on coliform and fecal coliform counts. Temperature of brine-salting had no effect on Enterobacteriaceae counts, but a significant effect of salting time on Enterobacteriaceae and fecal coliform counts was detected. Ripening temperature was the manufacturing variable with greatest influence (P<.01) on Enterobacteriaceae and coliform counts during the whole curing period.     

Influence of cooking and drying processes at different temperatures on the nutritive value of the protein of mussels (Mytilus edulis)

The effect of steam cooking (96 degrees C for 15 minutes) and drying at two temperatures, 70 degrees C and 110 degrees C, on nutritive value of mussel protein was studied. The measurements were carried out by nitrogen balance techniques in growing rats, and the nutritional parameters studied were: CD, BV and NPU. The crude digestibility (CD) values were: 87 +/- 1 and 82 +/- 1, and the biological values (BV), 80 +/- 1 and 74 +/- 1 for raw mussels, dried at 70 degrees C and at 110 degrees C respectively. This implies a significant decrease in the protein nutritive values of the mussel dried at a higher temperature. Cooking prior to drying significantly improved the digestibility and the biological value of the mussel's protein. In effect, improvement was so great, that the different drying temperatures did not affect the previously cooked product in a different way; therefore, the CD (94 +/- 1 and 94 +/- 1) and the BV (90 +/- 1 and 90 +/- 2) were the same for the mussel's protein, cooked and dried at 70 degrees C or at 110 degrees C.     

Extending the power line LAN up to the neighborhood transformer

This article reports on the performance of audio, video, multimedia, and other high-data-rate in-home networking applications. The article starts by describing the problems encountered in power line communication channels in terms of frequency response and noise characteristics, and explains how in-home power line LANs can be extended to the neighborhood transformer. OFDM physical layers providing speeds of 45 Mb/s and 200 Mb/s as well as QoS and security are introduced. Finally, the results of large tests involving several thousands of nodes are described.     

Lack of PPCA expression only partially coincides with lysosomal storage in galactosialidosis mice: indirect evidence for spatial requirement of the catalytic rather than the protective function of PPCA

Protective protein/cathepsin A (PPCA) is a pleiotropic lysosomal enzyme that complexes with beta-galactosidase and neuraminidase, and possesses serine carboxypeptidase activity. Its deficiency in man results in the neurodegenerative lysosomal storage disorder galactosialidosis (GS). The mouse model of this disease resembles the human early onset phenotype and results in severe nephropathy and ataxia. To understand better the pathophysiology of the disease, we compared the occurrence of lysosomal PPCA mRNA and protein in normal adult mouse tissues with the incidence of lysosomal storage in PPCA(-/-) mice. PPCA expression was markedly variable among different tissues. Most sites that produced both mRNA and protein at high levels in normal mice showed extensive and overt storage in the knockout mice. However, this correlation was not consistent as some cells that normally expressed high levels of PPCA were unaffected in their storage capability in the PPCA(-/-) mice. In addition, some normally low expressing cells accumulated large amounts of undegraded products in the GS mouse. This apparent discrepancy may reflect a requirement for the catalytic rather than the protective function of PPCA and/or the presence of cell-specific substrates in certain cell types. A detailed map showing the cellular distribution of PPCA in nomal mouse tissues as well as the sites of lysosomal storage in deficient mice is critical for accurate assessment of the effects of therapeutic interventions.     

Sj?gren syndrome and subacute demyelinating polyradiculopathy: an unusual association

INTRODUCTION: Sj?gren's syndrome is a chronic inflammatory condition of unknown aetiology and autoimmune pathology. The defining feature is the dry syndrome, expressed as xerophthalmia and xerostomia. Extra-glandular involvement at many other levels may also occur. Neurological involvement is not unusual. The peripheral nervous system is most frequently involved, and a predominantly sensitive symmetrical distal polyneuropathy may be the first sign of the condition. Other patterns of peripheral involvement are also associated with the syndrome. We present a case of subacute demyelinating polyradiculopathy associated with primary Sj?gren's syndrome. CLINICAL CASE: A 28 year old woman with dry syndrome presented with paraesthesia in her hands and feet, distal weakness, which had progressed proximally in the muscles of her arms and legs, and bilateral facial weakness. The condition progressed for eight weeks. When complementary tests were done, alterations typical of this condition (FR, ANA, anti-Ro and anti-La) were seen and also others typical of the dry syndrome (Schirmer's test). Therefore, in view of these findings and the clinical features, after other conditions had been ruled out, a diagnosis of primary Sj?gren's syndrome was made. The type of neuropathy was determined by the clinical features, electromyography and CSF findings. Treatment with corticosteroids gave good results. CONCLUSIONS: Demyelinating polyradiculopathy is a form of peripheral nervous system involvement which is rarely seen in this disorder. In the differential diagnosis Sj?gren's syndrome should be considered, an orientative history taken, autoantibodies determined and an ophthalmological examination made.     

A framework for avoiding steganography usage over HTTP

Steganographic techniques allow users to covertly transmit information, hiding the existence of the communication itself. These can be used in several scenarios ranging from evading censorship to discreetly extracting sensitive information from an organization. In this paper, we consider the problem of using steganography through a widely used network protocol (i.e. HTTP). We analyze the steganographic possibilities of HTTP, and propose an active warden model to hinder the usage of covert communication channels. Our framework is meant to be useful in many scenarios. It could be employed to ensure that malicious insiders are not able to use steganography to leak information outside an organization. Furthermore, our model could be used by web servers administrators to ensure that their services are not being abused, for example, as anonymous steganographic mailboxes. Our experiments show that steganographic contents can be successfully eliminated, but that dealing with high payload carriers such as large images may introduce notable delays in the communication process.