Tumor marker disaccharide D-Gal-beta 1, 3-GalNAc complexed to heat-labile enterotoxin from Escherichia coli.

Heat-labile enterotoxin (LT) is part of the cholera toxin (CT) family and consists of a catalytic A subunit and a B pentamer that serves to recognize the oligosaccharide part of the GM1 ganglioside receptor. We report here the crystal structure of heat-labile enterotoxin in complex with the disaccharide portion of the Thomsen-Friedenreich (T-antigen) tumor marker. The toxin:carbohydrate complex is determined to 2.13 A resolution, yielding an R-factor of 18.5%. The T-antigen disaccharide, D-Gal-beta 1,3-GalNAc-Ser/Thr, is present in more than 85% of human carcinomas and monitoring its autoimmune response is used for the early detection of tumors. Insight into the molecular recognition of this tumor antigen by sugar binding proteins can benefit the development of a diagnostic tool for human carcinomas as well as a T-antigen directed anticancer drug delivery system.     

Relation between stallion sperm binding to homologous hemizonae and fertility

The hemizona assay (HZA) has been developed as a diagnostic test to predict the fertilisation potential of human spermatozoa. The aim of this study was to develop an HZA for stallion spermatozoa and to investigate a possible relationship between fertility and the outcome of the HZA in this species. Equine oocytes were obtained from ovaries collected at a slaughterhouse and by transvaginal, ultrasound-guided follicle aspiration. They were then denuded from cumulus cells and stored in salt solution at 4 degrees C until use. On the day of the experiments the oocytes were bisected, thus providing 2 equal matching hemizonae from each oocyte. Semen samples from Dutch Warmblood stallions with known fertility data were used to assess the number of spermatozoa bound to the outer side of the hemizona after incubation in vitro. Sperm binding to matching hemizonae of a particular stallion was similar and confirmed the feasibility of using the HZA for the horse. Sperm hemizona binding capacity of 10 pairs of stallions was compared by incubating 1 hemizona with the semen of a stallion and the matching hemizona with the semen of another stallion from the same stud farm. Five matching pairs of hemizonae were used for each pair of stallions. There was a significant relationship between the mean number of spermatozoa bound to matching hemizonae and the fertility indices of stallions from each stud farm (P < 0.0001). It is concluded that HZA can be used as a valuable parameter in stallion semen analysis.     

Importance of dormancy and sink strength in sprouting of onions (Allium cepa) during storage

In onion ( Allium cepa L.) postponement of sprouting is necessary to achieve long term storage. We studied the factors determining sprouting during dry storage at 16°C. The period to visible sprouting depends on the length of the dormancy period, if present, and on the growth rate of the sprout. In the three cultivars tested, sprouts were initiated within 2 weeks after harvest indicating the absence of a real dormancy period. Sprout length increased linearly during storage. The mitotic activity of the apex decreased before harvest, was low at the transition from scale to leaf formation, and increased again when the sprout was initiated. From a few weeks before harvest, the initially high fructan content of the scales decreased, leading to a large increase in fructose. The sprout always contained enough carbohydrates for growth (between 50 and 60 mg g⁻¹ dry weight, of which 30% was fructan). The activity of sucrose synthase (EC 2.4.1.13) increased as the sprout grew, indicating an increase in sink strength. Invertase (EC 3.2.1.26) was absent in all bulb organs, during the various developmental stages. Although carbohydrates and enzymes were available for fast sprouting, sprout growth was still linear instead of exponential during dry storage at temperatures favorable for growth (16°C). The relative importance of factors determining sprouting are discussed.     

Soil nitrogen transformations and nitrate utilization by Deschampsia flexuosa (L.) Trin. at two contrasting heathland sites

Two Dutch heathland sites Hoorneboeg (HB) and Ede, dominated by Deschampsia flexuosa and differing in nitrate production, were sampled for an entire growing season. A large number of soil and plant parameters were monitored in an attempt to assess the contribution of nitrate in the N supply and its assimilation by Deschampsia.Average NO₃ ^– and NH₄ ⁺ concentrations (mg kg^–1) in the top 10-cm depth were 0.03 and 2.2, respectively, for HB, and 2.1 and 6.7, respectively, for Ede. Laboratory incubations of intact cores and experiments with FH-layer suspensions showed significantly higher mineralization and nitrification rates for the Ede site during most of the season. Nitrification was largely controlled by the rate of net N-mineralization, which in turn was highly affected by soil moisture. Nitrate production was virtually zero at HB and accounted for 25% of the net N-mineralization at Ede.Shoot chemical composition showed no essential differences for the two sites, but mean in vivo (current) foliar NRA was almost 2-fold higher at Ede than at HB, indicating some utilization of nitrate at the former location. At the HB site with essentially no nitrate production, however, enzyme activities were clearly higher than basal constitutive levels in NH₄ ⁺-fed plants. Apparently, shoot NRA at the HB site became positively affected by factors other than nitrate availability and/or showed disproportional increases in response to atmospheric nitrate inputs. Root NRA displayed the same low basal level at the two sites. Nitrate fertilization (100 kg N ha^–1) yielded maximally induced foliar NRAs similar to levels found in hydroponic nitrate plants. Although no accumulation of free NO₃ ^– was observed in shoots from fertilized plots, increases in foliar concentrations of both organic N and carboxylates clearly indicated nitrate assimilation. Root NRA showed no response to nitrate addition.It is concluded that current NRA measurements in Deschampsia at heathland sites are of limited value only, especially when interpreted in isolation. A combined approach, using concurrently conducted soil and plant analyses, will allow the extent of nitrate utilization in the field to be best characterized.Publication 2013 of the Netherlands Institute of Ecology.FAX no corresponding author: +31 8306 23227     

Mathematical simulation of the interactions among cyanobacteria, purple sulfur bacteria and chemotrophic sulfur bacteria in microbial mat communities

Abstract: A deterministic one-dimensional reaction diffusion model was constructed to simulate benthic stratification patterns and population dynamics of cyanobacteria, purple and colorless sulfur bacteria as found in marine microbial mats. The model involves the major biogeochemical processes of the sulfur cycle and includes growth metabolism and their kinetic parameters as described from laboratory experimentation. Hence, the metabolic production and consumption processes are coupled to population growth. The model is used to calculate benthic oxygen, sulfide and light profiles and to infer spatial relationships and interactions among the different populations. Furthermore, the model is used to explore the effect of different abiotic and biotic environmental parameters on the community structure. A strikingly clear pattern emerged of the interaction between purple and colorless sulfur bacteria: either colorless sulfur bacteria dominate or a coexistence is found of colorless and purple sulfur bacteria. The model predicts that purple sulfur bacteria only proliferate when the studied environmental parameters surpass well-defined threshold levels. However, once the appropriate conditions do occur, the purple sulfur bacteria are extremely successful as their biomass outweighs that of colorless sulfur bacteria by a factor of up to 17. The typical stratification pattern predicted closely resembles the often described bilayer communities which comprise a layer of purple sulfur bacteria below a cyanobacterial top-layer; colorless sulfur bacteria are predicted to sandwich in between both layers. The profiles of oxygen and sulfide shift on a diel basis similarly as observed in real systems.     

Aphid transmission of beet western yellows luteovirus requires the minor capsid read-through protein P74.

Beet western yellows luteovirus is obligately transmitted by the aphid Myzus persicae in a circulative, non-propagative fashion. Virus movement across the epithelial cells of the digestive tube into the hemocoel and from the hemocoel into the accessory salivary glands is believed to occur by receptor-mediated endocytosis and exocytosis. Virions contain two types of protein; the major 22 kDa capsid protein and the minor read-through protein, P74, which is composed of the major capsid protein fused by translational read-through to a long C-terminal extension called the read-through domain. Beet western yellows virus carrying various mutations in the read-through domain was tested for its ability to be transmitted to test plants by aphids fed on agro-infected plants and semi-purified or purified virus preparations. The results establish that the read-through domain carries determinants that are essential for aphid transmission. The findings also reveal that the read-through domain is important for accumulation of the virus in agro-infected plants.     

Fructosebisphosphatase isoenzymes of the chemoautotroph Xanthobacter flavus.

Xanthobacter flavus employs two fructosebisphosphatase (FBPase)-sedoheptulosebisphosphatase (SBPase) enzymes. One of these is constitutively expressed and has a high FBPase-to-SBPase ratio. The alternative enzyme, which is encoded by cbbF, is induced during autotrophic growth. The cbbF gene was expressed in Escherichia coli, and the FBPase was purified to homogeneity. The purified enzyme has a specific FBPase activity of 114 mumol/min/mg of protein, a Michaelis constant for fructosebisphosphate of 3 microM, and a low FBPase-to-SBPase ratio. CbbF was activated by ATP and inhibited by Ca2+.     

Transplantation of Foetal SCN into the Brain of non SCN-Lesioned Rats

Transplantation of foetal SCN tissue into the brain of arrhythmic SCN-lesioned rats and hamsters has shown to be effective in restoring circadian rhythms. Transplantation of the SCN into normal untreated rats has not been described so far as function is concerned. In rats certain demands have to be met for successful grafting of the SCN. Location, age and method of transplantation play an important role in the survival and function of the graft. This paper describes a method for SCN transplantation in normal rats. Results show transplant survival in 95% and successful grafting of SCN tissue in 85% of the treated rats as shown by VP and VIP staining. Disturbed circadian eating, drinking and activity rhythms are noted when grafts are located very near the endogenous SCN. Rhythms of wheel running and body temperature were less affected. The method described seems therefore well suited to do further research with circadian rhythms.