Knowledge-based modeling of the D-lactate dehydrogenase three-dimensional structure

A three-dimensional structure of the NAD-dependent D -lactate dehydrogenase of Lactobacillus bulgaricus is modeled using the structure of the formate dehydrogenase of Pseudomonas sp. as template. Both sequences share only 22% of identical residues. Regions for knowledge-based modeling are defined from the structurally conserved regions predicted by multiple alignment of a set of related protein sequences with low homology. The model of the D -LDH subunit shows, as for the formate dehydrogenase, an α/β structure, with a catalytic domain and a coenzyme binding domain. It points out the catalytic histidine (His-296) and supports the hypothetical catalytic mechanism. It also suggests that the other residues involved in the active site are Arg-235, possibly involved in the binding of the carboxyl group of the pyruvate, and Phe-299, a candidate for stabilizing the methyl group of the substrate. © 1995 Wiley-Liss, Inc.     

Biological,chemical and physical factors influencing the seasonal change of polysaccharide concentration in irrigation-channel sediment

Variation of polysaccharide concentration in irrigation-channel sediment was determined concurrently with biological, chemical and physical factors influencing the benthic algal community. Phenol-sulphuric acid method was used to measure polysaccharide concentration. Polysaccharide concentration, biomass of benthic algae, and species composition changed spatially and temporally. Fluctuations of total suspended solid (TSS) concentration and exposure of channel bed to direct sunlight had major effects on algal growth and polysaccharide production. Polysaccharide concentration was correlated to chlorophyll a concentration (r=0.73, P<0.001) and algal biomass (r=0.57, P<0.001). Fragilaria construens and Aulacoseira (Melosira) italica were the most common diatoms in the benthic flora. Chlorophyll a concentration in the sediment showed a strong negative correlation (r=-0.99, P<0.001) with the seasonal variation of TSS concentration in channel water. The polysaccharides produced by benthic microorganisms play a major role in clogging channel bed and thereby reducing seepage from earthen irrigation channels. Correlations between polysaccharide concentration and chlorophyll a (and algal biomass) further indicate the importance of benthic algae for polysaccharide production. Since availability of light to the algal flora is critical for the production of polysaccharides, the effect of clogging can be maximized by exposing the channel bed to direct sunlight during non-irrigation period (winter).     

DNA fingerprint analysis for the detection of induced mutations in mammalian cells in culture

A mutation assay in cultured mammalian cells based on the direct analysis of minisatellite DNA was developed. Band pattern variations reflect DNA alterations ranging from single base changes to complex rearrangements. By DNA fingerprinting a large number of autosomal loci throughout the human genome can be simultaneously checked, therefore minimizing the size of the samples of cell colonies to be scored in the absence of phenotypic selection. For the mutation assay chinese hamster cells (V79) were treated with Nitrosoguanidine and 14 independent colonies were isolated and expanded. DNA fingerprints were obtained after digestion of the DNA extracted from each clone with bothHinfI andHae III, and hybridisation with both 33.15 and 33.6 probes. Twelve colonies from untreated cells were also analysed. Several differences in the band pattern of treated colonies were observed when compared with untreated cells; digestion withHae III and hybridisation with 33.15 probe allowed the detection of the highest frequency of induced variants. The results suggest that minisatellite sequences are hypermutable sites that can be used to monitor the mutagenic potential of chemical agents directly at the DNA level, without phenotypic selection. Moreover, with the method herein decribed, it is possible to distinguish between true mutations and epimutations, such as those caused by changes in DNA methylation.     

IgM anti-hepatitis C virus in patients with chronic non-A, non-B hepatitis and their relationship to viral replication

Patients with hepatitis C virus (HCV) infection may have different patterns of antibody response to various structural and non-structural viral antigens. We have correlated the serological patterns to the clinical features of chronic infection and to viral replication in 68 HCV-Ab-positive patients with chronic liver disease at different stages (19 with cirrhosis-hepatocellular carcinoma, 38 with chronic active hepatitis and 11 with chronic persistent hepatitis). Serum samples from each patient were assayed for HCV-IgM by enzyme immunoassay and for HCV-RNA by the polymerase chain reaction using primer sets derived from the 5'-non-coding region. The prevalence of HCV-IgM was high (54 patients (79.4%)) and the study showed a good correlation between high values of anti-HCV-IgM and the presence of HCV-RNA in serum, since HCV-RNA was detected in 35 of the 54 IgM-positive patients (64.8%) and notably in 19 of the 20 subjects with high levels of specific IgM. Conversely, all the 35 sera containing HCV-RNA were also reactive for HCV-IgM, while none of the HCV-IgM-negative sera was HCV-RNA reactive. Positivity rates for both HCV-RNA and IgM anti-HCV were higher in the more advanced stages of disease; thus, the clinical pattern of HCV chronic hepatitis seems to be strictly related to the serological pattern and the presence of HCV-RNA.     

Lectin binding sites during Drosophila embryogenesis

The spectrum of lectin binding sites as it emerges during embryonic development of Drosophila was analysed by means of fluorescein-labelled lectins. As development and morphogenesis proceed, the reaction pattern becomes more and more complex. Mannose/glucose-, mannose-, N-acetylglucosamine- and poly-N-ace-tylglucosamine-specific lectins bind ubiquitously. Nuclear envelopes only have binding sites for wheat germ agglutinin. N-acetylgalactosamine-binding lectins are specific for ectodermal derivatives. Ga-3-N-acetylgalac-tosamine-binding lectins are highly selective markers for neural structures, haemocytes and Garland cells. It is also shown that Drosophila laminin is differentially glycosylated. The possible implications of differential and germ layer-specific glycosylation are discussed.Dedicated to the memory of Jan Callaerts     

Effects of metal speciation on growth of phytoplankton with special reference to iron

This paper is a compilation of studies concerning the effects of metal speciation on the growth of phytoplankton. Special attention is paid to the speciation and availability of iron in lake Tjeukemeer, The Netherlands. Under laboratory conditions the free ionic metal species generally appear to be most effective in determining metal availability and toxicity. A variety of factors controlling solubility, ion-exchange, complexation or chelation, sorption and electrostatical attraction of metal ions affect the metal speciation, mostly resulting in reduced availabilities. However, some organic metal chelates such as citrates, nitrillotriacetates and the specifically iron chelating siderophores, are sometimes more available than the corresponding free ions. The presence of other metals also influences the availability of a given metal by competing for the same binding sites on the algal cell. This competition leads to antagonism betweene.g. iron nutrition and cadmium toxicity in marine diatoms. In the eutrophic, alkaline, hard and humic lake Tjeukemeer, the free Fe³⁺ concentration is below measurable levels and does not match the iron requirement of the phytoplankton. So most of the algal iron must have been provided by the predominant inorganic iron colloids and particles bye.g. dissolution or photo-degradation (reduction). If the provision rates of available iron are slow in relation to that of iron uptake, the growth of some phytoplankton species may become iron-limited. Continuous culture work indicated that the iron fraction <0.2 m from Tjeukemeer,i.e., the soluble fraction, is about one third as much available as iron from NH₄Fe(SO₄)₂.12H₂O. Different phytoplankton species vary widely in their metal requirements and tolerances. Therefore, metal speciation and availability may affect species composition and succession within phytoplankton communities. So far the assessment of metal availability in natural waters has been complicated by the complex metal chemistry and by methodological limits.     

A comparative dynamic study of the effectiveness of gastric cytoprotection by vitamin A, De-Nol, sucralfate and ulcer healing by pirenzepine in patients with chronic gastric ulcer (a multiclinical and randomized study)

The gastric cytoprotective effects of vitamin A, De-Nol and sucralfate were compared with the effectiveness of pirenzepine in healing ulcer in patients with chronic gastric ulcer. A total of 100 patients was randomized into different groups: the patients were treated with antacids, vitamin A (3 X 50.000 IU), De-Nol liquid (4 X 5 ml), sucralfate (4 X 1 g) or pirenzepine (3 X 50 mg). The treatment was continued for 4 weeks. At the beginning, 2 and 4 weeks after starting treatment the patients were subjected to endoscopy and the size of the ulcer was measured planimetrically. The ulcer-healing effect of De-Nol liquid was significantly better than that of the antacids (p less than 0.01). Ulcer size was reduced significantly in all groups (p less than 0.01), however, at the end of the study the gastric ulcers were smallest in the De-Nol treated group (p less than 0.001). The dynamics of ulcer healing in the second week was most favourable in the patients receiving vitamin A (p less than 0.01). The present data point to the cytoprotective effects of De-Nol liquid, vitamin A and sucralfate and to their ability of healing chronic gastric ulcers.     

Studies on regulation of chorionic gonadotropin secretion in primates

The regulation of secretion of chorionic gonadotropin in primates has been studied using bothin vivo andin vitro models.In vivo studies using the pregnant bonnet monkey revealed that at the doses tested, the administration of progesterone or estradiol 17Β in combination or alone did not result in any appreciable change in the duration or magnitude of serum chorionic gonadotropin levels. However, administration of lutropin-releasing hormone by intravenous route resulted in significant increase in chorionic gonadotropin levels within 30–60 min and the extent of stimulation seemed to depend on the state of pregnancy. Forin vitro studies, explants or cells prepared from first trimester human placenta has been used. The functional integrity of these cells has been established by demonstrating the binding of [¹²⁵I]-labelled human chorionic gonadotropin antibody to the cells as well as the synthesis of [³H]-labelled human chorionic gonadotropin.In vitro studies using the cells revealed that addition of lutropin-releasing hormone caused a significant increase in chorionic gonadotropin and estradiol 17Β secreted into the medium. Thus bothin vivo andin vitro results suggest that lutropin-releasing hormone could be one of the factors involved in regulation of chorionic gonadotropin secretion in primates.