Characterization of peptides cleaved by plasmin from the C-terminal polymerization domain of human fibrinogen

The C-terminal region of the fibrinogen gamma chain is known to participate in several functional interactions including fibrin polymerization. This part of the molecule is retained on the gamma chain of fragment D (FgD) when fibrinogen is digested by plasmin in the presence of calcium to produce the fragment D-fragment E (FgD X FgE) complex but is lost if FgD is prepared in the absence of calcium. In an attempt to characterize the C-terminal polymerization domain we have used three techniques to examine this further degradation of FgD following the addition of EDTA and plasmin. Analysis of the digestion by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed a progressive cleavage of the gamma chain to two small remnants. The polymerization-inhibitory activity of the whole digest was studied using acid-solubilized fibrin. A progressive loss of inhibitory activity was associated with gamma chain shortening, reaching greater than a 120-fold reduction at the end of digestion. The cleavage of peptides was followed by reverse-phase high performance liquid chromatography and the release of a characteristic peptide triplet was associated with gamma chain cleavage. Manual sequencing, amino acid analysis, and fast atom bombardment mass spectrometry established the three peptides as gamma 303-356, 357-373, and 374-405. These peptides have sequences in common with those peptides recently reported by other investigators to be potent polymerization inhibitors. However, when a mixture of the three peptides was added in a 200-fold molar excess to polymerizing fibrin, no inhibitory activity could be demonstrated. It is concluded that the C-terminal polymerization domain of fibrinogen may be an extended region which includes the sequence gamma 303-405, when this is contiguous with the remainder of the gamma chain.     

Screening of white-rot fungi on (14C)lignin-labelled and (14C)whole-labelled wheat straw

Summary 74 Basidiomycetes have been tested for ligninolytic capability on (¹⁴C)lignin-labelled wheat straw. Fifteen strains were selected and rested more accurately for ligninolytic activity and the capacity to degrade wheat straw. The asymptote, inflexion point and degradation rate were determined using a model approach. The fungi exhibited very different responses with respect to lignin biodegradation: high asymptote for Pleurotus ostreatus (77%), low inflexion points for Sporotrichum pulverulentum Nov. (6.1 days) and Pycnoporus spp. (2.7 to 4.7 days) with high and slow degradation rates, respectively (0.91% and 0.45% of ¹⁴CO₂ release/day). Degradation values for (¹⁴C)whole-labelled wheat straw exhibited less variation. Finally, the strains Pleurotus ostreatus, Dichomitus squalens and Bjerkandera adusta showed the highest selectivity of lignin removal.     

A chromatin core particle obtained by selective cleavage of histones by clostripain.

Rat liver chromatin core particles digested with clostripain yield a structurally well-defined nucleoprotein particle with an octameric core made up of fragmented histone species (designated H'2A, H'2B, H'3 and H'4, respectively) after selective loss of a sequence segment located in the N-terminal region of each core histone. Sequential Edman degradation and carboxypeptidase digestion unambiguously establish that histones H2A, H2B, H3 and H4 are selectively cleaved at the carboxyl side of Arg 11, Lys 20, Arg 26 and Arg 19 respectively and that the C-terminal sequences remain unaffected. Despite the loss of the highly basic N-terminal regions, including approximately 17% of the total amino acids, the characteristic structural organization of the nucleosome core particle appears to be fully retained in the proteolyzed core particle, as judged by physicochemical and biochemical evidence. Binding of spermidine to native and proteolyzed core particles shows that DNA accessibility differs markedly in both structures. As expected the proteolyzed particle, which has lost all the in vivo acetylation sites, is not enzymatically acetylated, in contrast to the native particle. However, proteolyzed histones act as substrates of the acetyltransferase in the absence of DNA, as a consequence of the occurrence of potential acetylation sites in the core histones thus rendered accessible. The possible role of the histone N-terminal regions on chromatin structure and function is discussed in the light of the present observations with the new core particle obtained by clostripain proteolysis.     

Buspirone: A potential atypical neuroleptic

Buspirone produces a dose-dependent but short-lived elevation in striatal dopamine (DA) metabolites in the rat. $\text{In}$$\text{vitro}$, buspirone possesses an affinity similar to sulpiride for DA receptors (3H-spiperone). A moderate affinity for α₁ receptors was also observed while buspirone was inactive at α₂, β, muscarinic and serotonin₂ receptors. This pharmacological profile as well as previous behavioral data indicate that buspirone may be a potential “atypical” neuroleptic.     

A STUDY OF GLUCO-CORTICOSTEROID-INDUCED PYKNOSIS IN THE THYMUS AND LYMPH NODE OF THE ADRENALECTOMIZED RAT

Pyknotic nuclei, observed in the thymus of steroid-treated rats, are dense, homogeneous, intensely basophilic and Feulgen positive. Under the electron microscope, the image is that of a complete segregation of the chromatin from the nuclear sap producing a margin or crescent of condensed chromatin. Approximately 30% of all small thymocytes appeared to undergo this type of degeneration within 3–4 hr after administration of the synthetic corticosteroid, dexamethasone. At this time, pyknotic thymocytes were observed in clusters, probably as a result of the activity of dense reticular cells and macrophages. Topographical and experimental data suggest the existence of a select population of steroid-sensitive thymic cells. Furthermore, on the basis of thymidine-³H incorporation studies, it appears that the steroid-sensitive population of thymocytes does not correspond to "aged" cells. In addition, many plasma cells became pyknotic after the same steroid treatment, indicating an unexpected similarity between their nuclei and those of lymphocytes. Finally, steroid failed to induce pyknosis of thymocytes in a variety of in vitro experiments, suggesting that the in vivo effect of steroid is of an indirect nature. The results are discussed in terms of (a) the nature of the nuclear changes characterizing pyknosis, (b) the hypothetical mechanism whereby steroids trigger such changes, and (c) the population of cells susceptible to steroid-induced pyknosis.     

Analyse der schlagauslösenden Bewegungsparameter einer punktförmigen Beuteattrappe bei der Aeschnalarve

Zusammenfassung Bei der Libellenlarve Aeschna cyanea M. werden die einzelnen, für die Auslösung des Fangschlags relevanten Bewegungsparameter einer punktförmigen Beuteattrappe und die wirksamste Kombination dieser Parameter bestimmt.Als Beiz dient der beliebig bewegbare Leuchtpunkt eines Oszillographen, der auf die ebene Mattscheibe eines Versuchsaquariums projiziert wird. Die Schläge der frei beweglichen Larve werden vom Beobachter gezählt oder elektrisch registriert.Die Bahngeschwindigkeit von kontinuierlich gebotenen Bewegungsreizen wirkt sich stark auf die Schlagzahl aus: Die Schläge nehmen von 0,005–2,5 cm/sec zu, nehmen von 5,1 cm/sec an wieder ab und hören bei 41,0 cm/sec ganz auf. Die Veränderung der mittleren Geschwindigkeit von Sinusschwingungen und Zufallsbewegungen bewirkt ähnliche Reaktionskurven wie die Veränderung der gleichmäigen Geschwindigkeit von Dreiecksschwingungen und kreisförmigen Bewegungen; eine gleichförmige Optimalgesohwindigkeit wird jedoch stärker beantwortet als eine periodisch schwankende.Bietet man eine Folge von diskontinuierlichen Bewegungsreizen, die nach einer einmaligen Durchquerung eines begrenzten Feldes der Projektionsfläche verschwinden, so spielt die Dauer einer Einzelbewegung eine Rolle. Um die Schläge voll in Gang zu bringen, müssen eindimensionale Schwingungen 3–6 sec dauern, ein viel intensiver wirkender zweidimensionaler Reiz (Zickzackbewegung) jedoch nur 0,8 sec.Im optimalen, relativ hohen Geschwindigkeitsbereich läßt die Erhöhung der Bewegungsamplitude von 0,25 auf 2,0 cm die Schlagzahl progressiv absinken. Der Vergleich zwischen diesen Amplituden und dem Öffnungswinkel des frontalen, die Beute fixierenden Ommatidienfeldes zeigt, daß der Leuchtpunkt nur bei sehr kleinen Ablenkungen die frontalen Rezeptoren kontinuierlich reizt. — Die Bevorzugung von raschen Bewegungsreizen mit kleiner Amplitude besteht nicht bei Larven, die durch prompte Fixier- und Folgereaktionen den Leuchtpunkt in ihrer frontalen Fixierebene bewahren.Der Vergleich zwischen den 4 in dieser Untersuchung erzeugten Bewegungsmustern (ein- und zweidimensionale Schwingungen, Kreis- und Zufallsbewegungen) zeigt, daß eine Bewegung um so mehr Schläge auslöst, je vollständiger sie auf dicht aneinanderliegenden, zweidimensionalen Bahnen das frontale Ommatidienfeld abtastet.Die optimale Reizkombination (Zickzackbewegung) besteht aus einer kleinen (0,2–0,4 cm) Vertikalschwingung mit optimaler Geschwindigkeit, die sich langsam (0,32 cm/sec) seitlich verschiebt. Dieser Reiz stellt die Verbindung der optimalen Werte aller Bewegungsparameter dar und bewirkt, daß pro Zeiteinheit eine möglichst große Zahl frontaler Rezeptoren mit der optimalen Bahngeschwindigkeit gereizt wird.

---

Analysis of the parameters of a moving lightspot which release the predatory strike in dragonfly larvae

Summary This study analyses the predatory strike response of the dragonfly larva Aeschna cyanea M. towards a moving spot of light. Its aim is to determine the single parameters of movement of the spot which release the strike and the most effective combination of these parameters.As the velocity of a continuously moving lightspot is increased the number of strikes rises to a maximum (at 2.5 cm/sec) and then declines to 0 (at 41.0 cm/sec). Both uniform and non uniform velocities give curves of similar shape but different magnitudes.In the presentation of a sequence of discontinuous movements (where the spot moves across a part of the screen and then disappears) the duration of a single movement is important: Unidimensional oscillations must last 3 to 6 sec in order to release predatory strikes; twodimensional zigzag movements, much more effective, need last only 0.8 sec.In the optimal velocity range, increasing the amplitude of a movement from 0.25 to 2.00 cm produces a progressive decrease of the response rate. The comparison between these amplitudes and the size of the field of the frontal ommatidia, which fixate the prey, suggests that the spot stimulates these receptors continuously only when it moves with small amplitudes. — However, this preference for small amplitudes does not exist in those individuals which have rapid fixation- and following-reactions and which thus can track the stimulus.Comparison between the four patterns of movement which have been presented (one- and two-dimensional oscillations, circular and random movements) suggests that the spot releases more strikes the more exactly its movement covers the frontal fixation plane.The most effective stimulus for eliciting strikes was found to be a twodimensional zigzag oscillation. This movement consists of a small (0.2–0.4 cm), rapid (2.5 cm/ sec), vertical oscillation, which progresses slowly (0.3 cm/sec) sideways. This movement combines the optimal values of all parameters and stimulates with the optimal velocity the greatest possible number of frontal receptors in a given time.

---

---

Diese Arbeit wurde in Seewiesen mit Dr. H. C. Howland begonnen und dank der fortwährenden, großzügigen Unterstützung von Herrn Dr. H. Mittelstaedt beendet. Frau L. Dinnendahl fertigte die Zeichnungen an und durchsah das Manuskript, Dr. E. Kramer und Herr P. Heinecke standen mir ständig in technischen Fragen bei, Herr E. Butenandt leistete wertvolle Kritik am Manuskript. — In Genf gewährte mir Prof. J. Piaget vollkommene Freiheit in der Gestaltung meiner Arbeit. — Ihnen allen sei an dieser Stelle herzlichst gedankt.     

Electron Microscope Study of the Human Neuromuscular Junction

A preliminary electron microscope study of human neuromuscular junction is presented. The biopsy material was taken from the palmarus longus, and fixed routinely in osmium tetroxide and embedded in methacrylate. The structure of the motor endings and the relationship of the synaptic vesicles to the axolemmal membrane are described. The synaptic clefts are filled with an homogeneous material in continuity with the basement membrane covering the muscle fiber. The subneural apparatus is described, and special attention is paid to a vesicular component present in the sarcoplasm of the junctional area, which differs from synaptic vesicles and is presumed to be a derivate of the sarcoplasmic reticulum.     

Plasmid-mediated resistance to fosfomycin in Staphylococcus epidermidis

Staphylococcus epidermidis strain BM2641, isolated from a patient, was resistant to penicillin G, methicillin, aminoglycosides, chloramphenicol, macrolide, lincosamide and streptogramin B-type (MLS) antibiotics, and to high levels of fosmycin. Resistance to forsfomycin and/or to MLS was lost at low frequencies either spontaneously or after curing with novobiocin. The plasmid DNA from BM2641 and its cured derivatives was purified, analyzed by agarose gel electrophoresis and transferred to a nitrocellulose sheet. Comparative analysis of the resistance phenotypes with the plasmid content of the strains indicated that fosfomycin and MLS resistance were encoded by plasmids pIP1842 (2.5 kb) and pIP1843 (2.6 kb), respectively. Southern hybridization with a probe specific for gene fosA of Serratia marcescens showed that the fosfomycin resistance determinant in Staphylococcus is not homologous to that of Gram-negative bacteria.