Influence of sarmesin on the cardiac and vascular actions of angiotensin II

Summary The angiotensin II (ANG II) receptor blocker properties of sarmesin and its influence on the homotropic cooperativity of ANG II receptors were studied in two experimental models: isolated rabbit aorta and isolated rabbit atria. The results show that: (i) sarmesin is a specific competitive antagonist of ANG II receptors, with high affinity (pA₂=8.93 in the isolated aorta and 8.66 in the isolated atria); and (ii) the slope of the concentration-response curves to ANG II and the Hill coefficient increased in the presence of sarmesin, the latter suggesting an enhancement of the positive homotropic cooperativity of ANG II receptors. These results may be explained overall by the reciprocal negative modulation of receptor affinity between sarmesin and ANG II, due, possibly, to opposite effects on the binding of G-proteins to ANG II receptors.     

Optimized BlaM-transposon shuttle mutagenesis of Helicobacter pylori allows the identification of novel genetic loci involved in bacterial virulence

Helicobacter pylori is an important etiologic agent of gastroduodenal disease in humans. In this report, we describe a general genetic approach for the identification of genes encoding exported proteins in H. pylori. The novel TnMax9 mini-blaM transposon was used for insertion mutagenesis of a H. pylori gene library established in Escherichia coli. A total of 192 E. coli clones expressing active β-lactamase fusion proteins (BlaM⁺) were obtained, indicating that the corresponding target plasmids carry H. pylori genes encoding putative extracytoplasmic proteins. Natural transformation of H. pylori P1 or P12 using the 192 mutant plasmids resulted in 135 distinct H. pylori mutant strains (70%). Screening of the H. pylori collection of mutant strains allowed the identification of mutant strains impaired in motility, in natural transformation competence and in adherence to gastric epithelial cell lines. Motility mutants could be grouped into distinct classes: (i) mutant strains lacking the major flagellin subunit FlaA and intact flagella (class I); (ii) mutant strains with apparently normal flagella, but reduced motility (class II), and (iii) mutant strains with obviously normal flagella, but completely abolished motility (class III). Two independent mutations that exhibited defects in natural competence for genetic transformation mapped to different genetic loci. In addition, two independent mutant strains were isolated by their failure to bind to the human gastric carcinoma cell line Katoill. Both mutant strains carried a transposon in the same gene, 0.8 kb apart, and showed decreased autoagglutination when compared to the wild-type strain.     

High frequency electric fields for trapping of viruses

Summary Combining dielectrophoretic and hydrodynamic forces in micro electrode structures allows enrichment and stable trapping of viruses in aqueous solutions. Fluorescently labelled Influenza and Sendai viruses were collected from solutions of 2^*10⁵ – 2^*10⁸ viruses/l within a few seconds. In the central part of the trap a virus aggregate of about 2–9 m in diameter was formed. This corresponds to a local enrichment of viruses up to a factor of about 1400.     

β-Sheet Models for the Ordered Filamentous Structure Formed by a Peptide That Enhances the Action of Insulin

Certain peptides with sequences related to part of the major histocompatibility complex class I antigen enhance the action of insulin. These peptides also aggregate into fibrous structures that seem to be related to their biological activity. In the current study, the 17-residue peptide with amino acid sequence Gly-Asn-Glu-Gln-Ser-Phe-Arg-Val-Asp-Leu-Arg-Thr-Leu-Leu-Arg-Tyr-Ala is used as a representative example of these bioactive molecules. As seen by electron microscopy, the peptide associates into gently twisted ribbons, 50 Å thick, in which the amount of twist decreases as the ribbons become wider. X-ray diffraction analysis suggests that the peptides are arranged as in an antiparallel β-sheet extending essentially endlessly along the fiber axis. The amino acid sequence of the peptide is such that one side of the β-sheet is predominantly polar while the opposite side is nonpolar. This allows the β-sheets to form multilayers with alternating hydrophobic and hydrophilic interfaces. The length of the extended peptide (≈54 Å) determines the thickness of the ribbon and the tendency of individual β-sheets to twist accounts for the twisting of the ribbons. An alternative model is also discussed, again based on antiparallel β-sheets, but with adjacent sheets interdigitated in a “side-by-side” fashion rather than forming stacked layers. Comparable inactive peptides such as Gly-Asn-Glu-Gln-Ser- A_l_a_-Arg-Val-Asp-Leu-Arg-Thr-Leu-Leu-Arg-Tyr-T_y_r_ (changed amino acids underlined) do not form ordered filamentous structures.     

Gymnodinium catenatum Graham (Dinophyceae)in Europe: a growing problem?

The microreticulate resting cyst of the potentially toxic, chain-forming,unarmoured neritic dinoflagellate Gymnodinium catenalum Graham1943. the planktonic stage of which is not known from NorthEuropean waters, is reported for the first time from recentGerman coastal sediments of the North Sea and Baltic Sea. Insandy mud sediments of the German Bight, a maximum of 8 5 livingcysts cm^–3 were found. In Kiel Bight sediments G.catenalumwas found in maximum concentrations of 17.0 living cysts cm^–3.In surface waters of the German Bight resuspended G catenatumcysts were observed at concentrations of up to 3.6 cysts l^–1.Successful germination experiments conducted with natural seawatershow that the occurrence of a vegetative form of G.catenatumin northern Europe is very likely. The present study highlightsthat cyst surveys provide an important tool for the evaluationof areas with potential toxicity problems, as they may indicatethe presence of hitherto overlooked species in the water column.     

Altered Growth and Wood Characteristics in Transgenic Hybrid Aspen Expressing Agrobacterium tumefaciens T-DNA Indoleacetic Acid-Biosynthetic Genes

A key regulator of cambial growth is the plant hormone indoleacetic acid (IAA). Here we report on altered wood characteristics and growth patterns in transgenic hybrid aspen (Populus tremula L. x Populus tremuloides Michx.) expressing Agrobacterium tumefaciens T-DNA IAA-biosynthetic iaaM and iaaH genes. Eighteen lines simultaneously expressing both genes were regenerated. Of these, four lines, verified to be transgenic by northern blot analysis, were selected and raised under controlled growth conditions. All four lines were affected in their growth patterns, including alterations in height and stem diameter growth, internode elongation, leaf enlargement, and degree of apical dominance. Two transgenic lines, showing the most distinct phenotypic deviation from the wild type, were characterized in more detail for free and conjugated IAA levels and for wood characteristics. Both lines showed an altered IAA balance, particularly in mature leaves and roots where IAA levels were elevated. They also exhibited changes in wood anatomy, most notably a reduction in vessel size, an increase in vessel density, and changes in ray development. Thus, the recent development of techniques for gene transfer to forest trees enabled us to investigate the influence of an altered IAA balance on xylem development in an intact experimental system. In addition, the results demonstrate the possibility of manipulating wood properties in a forest tree through controlled changes of IAA concentration and distribution.     

Functional analysis of the iron-stress induced CP 43′ polypeptide of PS II in the cyanobacterium Synechococcus sp. PCC 7942

Under conditions of iron-stress, the Photosystem II associated chlorophyll a protein complex designated CP 43, which is encoded by the isiA gene, becomes the major pigment-protein complex in Synechococcus sp. PCC 7942. The isiB gene, which is located immediately downstream of isiA, encodes the protein flavodoxin, which can functionally replace ferredoxin under conditions of iron stress. We have constructed two cyanobacterial insertion mutants which are lacking (i) the CP 43 apoprotein (designated isiA ^–) and (ii) flavodoxin (designated isiB ^–). The function of CP 43 was studied by comparing the cell characteristics, PS II functional absorption cross-sections and Chl a fluorescence parameters from the wild-type, isiA ^– and isiB ^– strains grown under iron-stressed conditions. In all strains grown under iron deprivation, the cell number doubling time was maintained despite marked changes in pigment composition and other cell characteristics. This indicates that iron-starved cells remained viable and that their altered phenotype suggests an adequate acclimation to low iron even in absence of CP 43 and/or flavodoxin. Under both iron conditions, no differences were detected between the three strains in the functional absorption crossection of PS II determined from single turnover flash saturation curves of Chl a fluorescence. This demonstrates that CP 43 is not part of the functional light-harvesting antenna for PS II. In the wild-type and the isiB ^– strain grown under iron-deficient conditions, CP 43 was present in the thylakoid membrane as an uncoupled Chl-protein complex. This was indicated by (1) an increase of the yield of prompt Chl a fluorescence (F_o) and (2) the persistence after PS II trap closure of a fast fluorescence decay component showing a maximum at 685 nm.Abbreviations Chl chlorophyll - CP 43, CP 47 and CP 43 Chl a binding protein complexes of indicated molecular mass - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - F_m and F_m fluorescence when all PS II reaction centers are dosed in dark- and light-acclimated cells, respectively - F_o fluorescence when all PS II reaction centers are open in dark acclimated cells - F_v variable fluorescence after dark acclimation (F_m–F_o)