Theoretical study of the reduction of nitric oxide in an A-type flavoprotein

The mechanism for the reduction of nitric oxide to nitrous oxide and water in an A-type flavoprotein (FprA) in Moorella thermoacetica, which has been proposed to be a scavenging type of nitric oxide reductase, has been investigated using density functional theory (B3LYP). A dinitrosyl complex, [{FeNO}⁷]₂, has previously been proposed to be a key intermediate in the NO reduction catalyzed by FprA. The electrons and protons involved in the reduction were suggested to “super-reduce” the dinitrosyl intermediate to [{FeNO}⁸]₂ or the corresponding diprotonated form, [{FeNO(H)}⁸]₂. In this type of mechanism the electron and/or proton transfers will be a part of the rate-determining step. In the present study, on the other hand, a reaction mechanism is suggested in which N₂O can be formed before the protons and electrons enter the catalytic cycle. One of the irons in the diiron center is used to stabilize the formation of a hyponitrite dianion, instead of binding a second NO. Cleaving the N–O bond in the hyponitrite dianion intermediate is the rate-determining step in the proposed reaction mechanism. The barrier of 16.5 kcal mol⁻¹ is in good agreement with the barrier height of the experimental rate-determining step of 14.8 kcal mol⁻¹. The energetics of some intermediates in the “super-reduction” mechanism and the mechanism proceeding via a hyponitrite dianion are compared, favoring the latter. It is also discussed how to experimentally discriminate between the two mechanisms. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Type I Restriction–Modification Loci Reveal High Allelic Diversity in Clinical Helicobacter pylori Isolates

Background: A remarkable variety of restriction‐modification (R‐M) systems is found in Helicobacter pylori. Since they encompass a large portion of the strain‐specific H. pylori genes and therefore contribute to genetic variability, they are suggested to have an impact on disease outcome. Type I R‐M systems comprise three different subunits and are the most complex of the three types of R‐M systems. Aims: We investigated the genetic diversity and distribution of type I R‐M systems in clinical isolates of H. pylori. Material and methods: Sixty‐one H. pylori isolates from a Swedish hospital based case‐control study and 6 H. pylori isolates of a Swedish population‐based study were analyzed using polymerase chain reaction for the presence of the three R‐M systems' subunits. Representative gene variants were sequenced. Results: Although the hsdM and hsdR genes appeared conserved in our clinical H. pylori isolates, the sequences of the hsdS loci were highly variable. Despite their sequence diversity, the genes per se were present at high frequencies. We identified a number of novel allelic hsdS variants, which are distinct from corresponding hsdS loci in the sequenced H. pylori strains 26695, J99 and HPAG1. In analyses of paired H. pylori isolates, obtained from the same individuals with a 4‐year interval, we observed genetic modifications of hsdS genes in patients with atrophic gastric mucosa. Discussion: We propose that the genetic variability of hsdS genes in a bacterial population will give rise to new specificities of these enzymes, which might lead to adaptation to an ever‐changing gastric environment.     

Soil seed bank and species turnover in a limestone grassland

Abstract. The vegetation and soil seed bank in a limestone grassland in southern Sweden were studied in permanent 1-m² subplots which had been either grazed or not grazed for 17 yr. Of the 92 species recorded, 18 were present only in the seed bank, 28 were more frequent and 24 were less frequent in the seed bank than in the vegetation and 22 were not detected in the seed bank. Among the species present in the seed bank, therophytes were over-represented. Species turnover in the vegetation was estimated from presence/absence data collected in 1980, 1986 and 1990. Turnover was high, but there were no differences between grazed and ungrazed subplots. The turnover for individual species was also high in many cases. There was no clear relationship between the turnover of a species and its presence in the persistent seed bank.     

Two novel IgG endopeptidases of Streptococcus equi

Streptococcus equi ssp. equi causes strangles, a highly contagious and serious disease in the upper respiratory tract of horses. Streptococcus equi ssp. zooepidemicus , another subspecies of this genus, is regarded as an opportunistic commensal in horses. The present study describes the characterization of two novel immunoglobulin G (IgG) endopeptidases of these subspecies, IdeE2 and IdeZ2. Both enzymes display sequence similarities with two previously characterized IgG endopeptidases, IdeE of S. equi ssp. equi and IdeZ of S. equi ssp. zooepidemicus . IdeE2 and IdeZ2 display high substrate-specificity in comparison with IdeE and IdeZ, as they both completely cleave horse IgG, while the activity against IgG from mouse, rabbit, cat, cow, sheep and goat is low or absent. The potential use of IdeE and IdeE2 as vaccine components was studied in a mouse infection model. In this vaccination and challenge study, both enzymes induced protection against S. equi ssp. equi infection.     

Food availability and nutrition in a seasonal perspective: A study from the Rukwa region in Tanzania

Most of the studies on seasonality in food supply and nutritional status have been carried out in areas characterized by extreme climatic conditions. This study was conducted in an area where the climate is favorable for grain cultivation. However, a large part of the population was found to face seasonal variations in food availability, most critically three to four months before the main harvest. Women lost about 3 percent of their weight in this period compared to the post harvest period. The nutritional status of preschool children declined in the lower socioeconomic group. These changes also coincided with a period of hard agricultural work, especially for women. The paper deals with factors affecting the duration of the maize stocks as well as strategies used when maize stocks are exhausted.     

Effect of cibacron blue on tubulin assembly in the absence and presence of microtubule-associated proteins

Cibacron blue was found to inhibit assembly and increase the critical concentration of microtubule proteins. In the presence of 4 mol Cibacron blue/mol tubulin, assembly was completely inhibited and pre-formed microtubules disassembled. Addition of 8% (v/v) dimethylsulfoxide to Cibacron blue-inhibited samples induced assembly of normal microtubules in addition to sheets of protofilaments. Disassembly was induced upon addition of 1 mM colchicine or 2mM Ca²⁺. No obvious difference was seen in the protein composition of these microtubules compared with controls. GTP exchange was not affected by the presence of Cibacron blue nor was GTP able to counteract its effect. This indicates that the exchangeable GTP site is not involved. The extent of assembly of phosphocellulose purified tubulin in the presence of 8% (v/v) dimethylsulfoxide was only slightly less in the presence of Cibacron blue, although the assembly rate was decreased. These results suggest that Cibacron blue might alter the binding of one or more of the associated proteins stimulating assembly.     

Effects of growth conditions on superoxide dismutase and catalase activities inSaccharomyces cerevisiae var.ellipsoideus

The superoxide dismutase (SOD) activity inSaccharomyces cerevisiae var.ellipsoideus increased when the cells were exposed to hyperbaric oxygen tension. Ethanol-grown cells, having a more oxidative metabolism, showed higher SOD activities than did glucosegrown cells. In a glucose-limited chemostat the SOD activity increased with the specific oxygen consumption rate. The increase in SOD activity may be explained by a higher intracellular flow of superoxide radicals at higher respiration rates. The catalase activity decreased with increasing growth rates in a glucose-limited chemostat, and the activity was lower in glucosethan in ethanol-grown cells.