Fluoritisierte Radiolarien aus Kieselkalk-Bänken des Mittel-Viseums (Unterkarbon) des Rheinischen Schiefergebirges (Deutschland)

From siliceous shales (Lower Carboniferous, Rheinisches Schiefergebirge), in direct neighborhood to a bed of allodapic limestone, the following fluontized radiolarian fauna has been extracted by chemical transformation of originally calcified skeletons:Albaillella cartalla, Latentifistula turgida, Eostylodktya cf. eccentrica, Tetragregnon sycamorensis sycamorensis, Belowea variabilis, Callela? hexactinia, Entactinia tortispina and Entactinia variospina. The limestone bed has been dated by calcareous foraminifera as being mid Visean in age (V 2b-3a, Cf 5-foraminiferal zone). The diagenetic calcification took place after the selective dissolution of the skeletons and was in itself not selective.     

Diurnal periodicity of chalcone-synthase activity during the development of oat primary leaves

Chalcone-synthase (CHS) activity was followed during the development of primary leaves of oat (Avena sativa L.) seedlings grown under different illumination conditions. Continuous darkness and continuous light resulted in similar time courses of enzyme activity. The maximum of CHS activity in etiolated leaves was delayed by 1 d and reached about half the level of that of light-grown leaves. In seedlings grown under defined light-dark cycles a diurnal rhythm of CHS activity and its protein level was observed which followed the rhythm of CHS-mRNA translational activity (Knogge et al. 1986). This rhythm persisted in continuous light after a short-term pre-exposure to the light-dark cycle but not in continuous darkness.Abbreviations CHS chalcone synthase - PAL phenylalanine ammonio lyase Financial support by the Deutsche Forschungsgemeinschaft is gratefully acknowledged (G.W., We 630/9-7; We 630/10-1). Thanks are given to Dr. St. Kellam (Department of Plant Microbiological Sciences, University of Canterbury, New Zealand) for correcting the English.     

Specific eradication of micrometastases by transfer of tumour-immune T cells from major-histocompatibility-complex congenic mice

Summary DBA/2 (H-2^d) mice bearing a transplanted highly metastatic lymphoma (ESb) in a state of widely disseminated disease could be successfully treated by a combination of surgery (removal of the local tumour), irradiation (5 Gy) and adoptive immunotherapy. The immunotherapy was achieved by transfer of anti-ESb-immune spleen cells from B10.D2 mice, which express the same major histocompatibility complex (MHC) molecules as DBA/2. In contrast, anti-ESb-immune cells from MHC-disparate C57BL/6 mice did not confer protective immunity. The B10.D2 anti-ESb-immune T cells contain two types of cytolytic specificity as detected by limiting-dilution analysis: (1) clones with specificity for the ESb-tumour-associated transplantation antigen (TATA) (at low frequency), and (b) clones with specificity for minor DBA/2 histocompatibility (H) antigens (at high frequency). Immune B10.D2 cells raised against different tumour lines or against TATA^– ESb tumour variants did not confer the 100% protection seen with immune cells against ESb TATA⁺ cells. Finally we demonstrate that the allogeneic immune cells are more potent in terms of protective immunity than corresponding syngeneic immune cells. The data suggest that the strong graft-versus-leukemia effect with immune T cells from allogeneic MHC-identical but not from MHC-disparate mice was due to T cells with MHC-restricted specificity for an ESb-associated TATA. A graft-versus-host reactivity that developed much later and could not be prevented was most likely due to T cells sensitized against normal minor H antigens of the host. Our results are of potential relevance for allogeneic bone marrow transplantation and adoptive immunotherapy protocols.     

1H nuclear-magnetic-resonance studies of the three-dimensional structure of the cardiotoxin CTXIIb from Naja mossambica mossambica in aqueous solution and comparison with the crystal structures of homologous toxins

Using the previously reported sequence-specific 1H-NMR assignments, structural constraints for the cardiotoxin CTXIIb from Naja mossambica mossambica were collected. These include distance constraints from nuclear Overhauser enhancement measurements both in the laboratory and in the rotating frame, dihedral angle constraints derived from spin-spin coupling constants, and constraints from hydrogen bonds and disulfide bridges. Structure calculations with the distance geometry program DISMAN confirmed the presence of the previously identified antiparallel beta-sheets formed by residues 1-5 and 10-14, and by 20-27, 35-39 and 49-55, and established the nature of the connections between the individual beta-strands. These include a right-handed crossover between the two peripheral strands in the triple-stranded beta-sheet, and a type I tight turn immediately preceding the beta-strand 49-55. The spatial arrangement of the polypeptide backbone in the solution structure of CTXIIb is closely similar to that in the crystal structure of the homologous cardiotoxin VII4 from the same species. In an Appendix the origin of the large pH dependence of two amide proton chemical shifts in CTXIIb is explained.     

Molecular characterization of the hemolysin determinant of Serratia marcescens.

The nucleotide sequence of a 7.3-kilobase-pair fragment of DNA encoding a hemolytic activity from Serratia marcescens was determined. Two large open reading frames were identified, designated shlA (Serratia hemolysin) and shlB, capable of encoding polypeptides of 165, 056 and 61,897 molecular weight, respectively. Both reading frames were expressed in vivo. The shlB gene product was localized to the outer membrane of Escherichia coli cells harboring the S. marcescens hemolysin determinant. Consistent with this location, a signallike sequence was identified at the N terminus of the polypeptide predicted from the nucleotide sequence of the shlB gene. Hyperexpression of the shlB locus permitted the identification of two shlB-encoded polypeptides of 65,000 and 62,000 molecular weight, respectively. Determination of the N-terminal amino acid sequence of the purified 62,000-molecular-weight protein confirmed that it was the mature form of the ShlB protein initially synthesized as a precursor (65,000-molecular-weight protein). By using polyclonal antisera raised against the purified proteins, ShlA and ShlB were identified in the outer membrane of S. marcescens. The shlA gene product was shown to interact with erythrocyte membranes, confirming it as the hemolysin proper. Both hemolysis and the interaction of ShlA with erythrocyte membranes did, however, require the ShlB function. Progressive deletion of the C terminus of the ShlA protein gradually reduced hemolytic activity until 37% of the amino acids had been removed. Elimination of 54% of the amino acids produced a nonhemolytic protein which, however, was still capable of associating with erythrocyte membranes.     

Enzymatic properties of proteolytic derivatives of human alpha-thrombin

The use of derivatives of alpha-thrombin obtained by limited proteolysis, that have only a single peptide bond cleaved, allowed the unequivocal correlation between the change in covalent structure and alteration of the enzymatic properties. beta T-Thrombin contains a single cleavage in the surface loop corresponding to residues 65-83 of alpha-chymotrypsin [Birktoft, J. J., & Blow, D. M. (1972) J. Mol. Biol. 68, 187-240]. Compared with alpha-thrombin, this modification had a minor effect on the following: (1) The Michaelis constant (Km) for two tripeptidyl p-nitroanilide substrates increased 2-3 fold, whereas the catalytic constant (k cat) remained unaltered. (2) A 2-3 fold increase in the binding constant (KI) of a tripeptidyl chloromethane inhibitor was observed, but the inactivation rate constant (k i) was the same, which indicated that the nucleophilicity of the active-site histidyl residue had not changed. (3) The second-order rate constant for the inhibition by antithrombin III decreased 2-fold. Heparin accelerated the inactivation, and the degree of acceleration was similar to that obtained with alpha-thrombin. Pronounced effects of the cleavage of this loop were found. (1) The cleavage of fibrinogen was approximately 80-fold slower than that with alpha-thrombin. This was mainly due to a 40-fold decrease in k cat. In contrast, only a 1.9-fold increase in the Michaelis constant was observed. (2) The affinity for thrombomodulin had decreased 39-fold compared to alpha-thrombin. epsilon-Thrombin contains a single cleaved peptide bond in the loop corresponding to residues 146-150 in alpha-chymotrypsin.(ABSTRACT TRUNCATED AT 250 WORDS)     

Analysis of outer membrane proteins which are associated with growth of Bacteroides thetaiotaomicron on chondroitin sulfate.

By analyzing outer membrane proteins of Bacteroides thetaiotaomicron on two-dimensional polyacrylamide gels, we were able to identify 10 protein spots that were associated with growth on chondroitin sulfate but not with growth on glucuronic acid or other monosaccharides. These proteins were distinct from the outer membrane polypeptides that were associated with growth on two other negatively charged polysaccharides, polygalacturonic acid and heparin. Of the 10 protein spots that were associated with growth on chondroitin sulfate, 4 could be detected on immunoblots with antiserum that had been raised against outer membranes from bacteria grown on chondroitin sulfate and then cross-adsorbed with membranes from bacteria grown on glucose. Synthesis of these four proteins appeared to be regulated coordinately with synthesis of the two enzymes that degrade chondroitin sulfate, chondroitin lyase I and II. Although one of the four proteins (Mr 110,000) was similar in molecular weight to the chondroitin lyases, the cross-adsorbed antiserum which detected this outer membrane protein did not cross-react with either of these two enzymes.     

Identification and genetic mapping of a herpes simplex virus capsid protein that binds DNA

Herpes simplex virus virion protein 19C (VP19C) is a constituent of both unenveloped (nuclear) and enveloped (cytoplasmic) capsids. In this paper we report that 32P-labeled DNA, either supercoiled or linear double stranded, efficiently bound to VP19C electrically transferred from denaturing polyacrylamide gels containing electrophoretically separated proteins from purified capsids. Analyses of the polypeptides specified by herpes simplex virus type 1 X herpes simplex type 2 recombinants with respect to electrophoretic mobility and binding of 32P-labeled DNA indicate that VP19C maps at the same location as infected cell polypeptide 32 and is derived from it.