Solid-state modifications of poly(γ-ethyl-L-glutamate)

Several modification of the arrangements of α-helical molecules were found in the solid films of poly (γ-ethyl-L -glutamate), depending on the casting solvent and the temperature. The helical conformation is somewhat looser than the normal 18-residue, 5-turn α-helix. Using x-ray diffraction, the types of molecular arrangements were classified into tetragonal, pseudohexagonal, and hexagonal ones. Tetragonal packing was observed in the filmm (form T) prepared by casting the solution in trifluorethanol or dichlorethane. The sample obtained from chloroform solution is a well-ordered, pseudohexagonal modification (form I). Forms I and T change into a poorly crystalline form III by annealing at temperatures above 130° C. It is particularly noteworthy that the less-ordered form III exhibits a thermoreversible transition around 110°C into a well-ordered form H with the hexagonal molecular packing.     

Polyclonal Antibody Production in Murine Spleen Cells Induced by Staphylococcus

Polyclonal plaque-forming cell (PFC) responses in murine spleen cells induced by Staphylococcus aureus and S. epidermidis were studied. Injection of Balb/c mice with S. aureus strain 248βH resulted in the generation of anti-trinitrophenyl (TNP) and anti-sheep red blood cell PFC in their spleens. Cultures of Balb/c spleen cells in the presence of S. aureus 248βH, Cowan I, or a protein A-deficient mutant yielded many anti-TNP PFC. The larger the number of organisms that were added to the cultures, the better was the PFC response. Both living and killed organisms, were capable of inducing the response, but an excess of living 248βH organisms in the cultures abrogated the response. All of the organisms (12 strains of S. aureus and 11 strains of S. epidermidis) freshly isolated from patients had the ability to induce the polyclonal PFC response in cell cultures. These organisms stimulated cultured C3H/HeJ mouse spleen cells, which were unresponsive to bacterial lipopolysaccharide (LPS). Cultured cells from the spleens of athymic nu/nu mice also responded to these organisms, and the number of PFC in nu/nu cell cultures was always greater than that in nu/+ cells prepared from a haired litter mate. Moreover, the responses of nu/nu spleen cell cultures to which nylon wool column-filtered splenic nu/+ T cells were added were lower than expected. These findings suggest that the polyclonal PFC response to staphylococci is thymus independent, but that the magnitude of the response is regulated by mature T cells. Cultures of macrophage-depleted spleen cells responded to the organisms to an extent similar to that of the control. The 248βH organisms were less capable of stimulating spleen cells of 2-week-old mice (i.e., early maturing B cells) than LPS. However, spleen cells from adult (7-week-old) and aged (9-month-old) mice responded well to both the organisms and LPS. Previous sensitization with the organisms in vivo did not affect any polyclonal responses of spleen cells in vitro to either the organisms or LPS. The role of staphylococcal protein A in the polyclonal PFC response to staphylococci is discussed.     

Settling-site selection and survival of two scale insects,Ceroplastes rubens andC. ceriferus,on citrus trees

We studied settling-site selection and the resulting survival of two sessile scale insects, Ceroplastes rubens and C. ceriferus, in the citrus tree, Citrus unshiu, in central Japan. C. rubens preferred 0-year-old twigs most as a settling-site; the density of nymphs settling on 0-year-old twigs was significantly higher than those on ≥1-year-old twigs, and few nymphs settled on ≥3-year-old twigs. The mean survival rates from settling until reproduction in the next year were significantly higher on more preferred twigs than on less preferred ones. In C. ceriferus, nymphs significantly preferred 1- and 2-year-old twigs to 0- and ≥3-year-old ones, and the mean survival rates on the more preferred 1- and 2-year-old twigs were significantly higher than those on less preferred ≥3-year-old twigs. However, the survival rate on less preferred 0-year-old twigs was slightly higher than those on 1- and 2-year-old ones. Thus, in both species of scale, it was the preferred twigs which were more profitable sites for survival after settling, except for less preferred 0-year-old twigs for C. ceriferus. In both scale species, most mortality was due to growth cessation, which is believed to be related to the twig quality as a food source. Predators and parasitoids were minor mortality factors. Both species showed constant survival rates until the density of settled nymphs exceeded double the “upper-limit” density, whereupon they decreased drastically. Nymphs of C. rubens settling on twigs of high scale density showed a spacing-out distribution, those of C. ceriferus did not. In C. rubens, an increase in preference for originally less profitable twigs at the later stage of the settling season was observed, but not in C. ceriferus. Accordingly, individuals of C. rubens showed a stronger tendency to avoid conspecifics than did C. ceriferus. Although nymphs of the two scales clearly preferred more profitable sites, their settling-site selection did not agree with the predictions from the ideal free distribution theory (Fretwell and Lucas, 1970). The discrepancies were (1) frequent settling on less profitable sites at the early stage of the settling season, (2) insufficient utilization of the most profitable twigs, and (3) virtually 100% mortality on overcrowded twigs under conditions where unoccupied profitable twigs still remained. These discrepancies are thought due to the limited dispersal time of nymphs.     

Application of leaf extract causes repetitive action potentials inBiophytum sensitivum

A wound stimulus evoked a number of repetivite action potentials in the leaf ofBiophytum sensitivum. When the cut end of a leaf was immersed in a leaf extract, the resulting repetitive action potentials continue for a long time. These repetitive action potentials disappeared immediately when the leaf extract, which contains a proposed stimulant, was removed and the cut end was washed with water.     

Rhythm in potassium uptake by a duckweed, Lemna gibba G3

The potassium uptake activity of the "flow-medium culture" ofa long-day duckweed, Lemna gibba G3, followed a circadian rhythmwhich persisted for more than 5 days under continuous light.The period of the rhythm was about 25 hr under 3000 lux at 26?Cand was slightly over-compensated against temperature, Q₁₀ beinga little less than 1.0. The amplitude of the rhythm was dependenton light intensity, and there was no potassium uptake in thedark. Magnesium uptake was affected by the potassium movementand showed circadian rhythmicity with a small amplitude underconditions where the potassium uptake was already saturated.Calcium uptake did not show any obvious rhythm. In Contrastto L. gibba, a short-day duckweed L. perpusilla 6746 displayedcircadian rhythm of potassium uptake only in the dark and notin the light. This rhythm did not persist beyond the secondcycle. (Received June 13, 1978; )     

Role of prostaglandin endoperoxides in the serum thiobarbituric acid reaction

In the presence of heme and reduced glutathione, prostaglandin (PG) endoperoxides underwent rapid conversion to malondialdehyde and 12l-hydroxy-5,8,10-heptadecatrienoic acid. In addition, PG endoperoxides as well as lipid peroxides produced malondialdehyde to yield a red pigment during the thiobarbituric acid reaction with different efficiencies. The relative rates of the reaction were: 1,1,3,3-tetraethoxypropane, 100; PGG₂, 55; PGH₂, 32; and 15-hydroperoxyarachidonic acid, 6. The thiobarbituric acid reactive materials in rabbit serum decreased by 25–60%, after intravenous administration of aspirin (a cyclo-oxygenase inhibitor) and with a concomitant decline of serum PG levels. These results, taken together, suggested that serum thiobarbituric acid values, considered to be an indicator of lipid peroxide levels, were to a significant extent due to PG endoperoxides and their derivatives.     

Ionic composition and pH of the vacuolar sap in marine dinoflagellate Noctiluca

Ionic composition of the vacuolar sap of Noctiluca miliariswas as follows: [Na⁺] = 487.3 mM, [K⁺]=24.1 mM, [Ca²⁺]=6.6 mM,[Mg²⁺]=2.8 mM, [Cl^–]=500mM, [NH₄⁺]=15–25 mM, and[SO₄^2–]=undetectable. To measure the vacuolar pH of singleliving cells, a pH-sensitive glass microelectrode was used.The vacuolar pH value was 3.50 ±0.18. When the cellswere transferred from normal sea water into osmotically adjusted50% sea water for one day, the vacuolar ion concentrations remainedalmost constant. Upon immersing the cells in osmotically unadjustedsea water of various concentrations for one day, the observedincrements or decrements of the vacuolar ion concentrationscould be accounted for largely by the migration of water outof or into the cells. The intrinsic ionic composition of thevacuole seems to be constant against changes in ion concentrationsof the bathing medium. (Received October 20, 1975; )     

Identification of nucleobase chemical modifications that reduce the hepatotoxicity of gapmer antisense oligonucleotides

Currently, gapmer antisense oligonucleotide (ASO) therapeutics are under clinical development for the treatment of various diseases, including previously intractable human disorders; however, they have the potential to induce hepatotoxicity. Although several groups have reported the reduced hepatotoxicity of gapmer ASOs following chemical modifications of sugar residues or internucleotide linkages, only few studies have described nucleobase modifications to reduce hepatotoxicity. In this study, we introduced single or multiple combinations of 17 nucleobase derivatives, including four novel derivatives, into hepatotoxic locked nucleic acid gapmer ASOs and examined their effects on hepatotoxicity. The results demonstrated successful identification of chemical modifications that strongly reduced the hepatotoxicity of gapmer ASOs. This approach expands the ability to design gapmer ASOs with optimal therapeutic profiles.     

Cooperative conformational change and aggregation of concanavalin A in alkaline solutions

Three properties, the binding activity to Sephadex G-75, conformation, and the extent of aggregation, of concanvalin A. (con A) in alkaline pH solutions were examined with special attention to the time course and their time-independent final values. Highly cooperative conformational changes among four subunits were suggested which were coupled either with protonation in the case of demetallized con A or with metal binding in the case of metal-liganded con A. Midpoints of the conversions of the metal-liganded con A were about pH 8.8, 9.1 and 9.1 with respect to the activity, the conformational change and the aggregation, respectively. These values were about 1 pH higher than the corresponding values of demetallized con A: 7.9, 8.05 and 8.2. Each conversion took place in narrow pH ranges. The pH range for the loss of activity was found to be significantly lower than those of the other two. The aggregation was suggested not to be coupled with the conformational change. Dissociation into subunits did not take place indicating strong interactions among four subunits in the tetramer.     

Purification and cDNA cloning of a cellulase from abalone Haliotis discus hannai.

A cellulase [endo-beta-1,4-D-glucanase (EC 3.2.1.4)] was isolated from the hepatopancreas of abalone Haliotis discus hannai by successive chromatographies on TOYOPEARL CM-650M, hydroxyapatite and Sephacryl S-200 HR. The molecular mass of the cellulase was estimated to be 66 000 Da by SDS/PAGE, thus the enzyme was named HdEG66. The hydrolytic activity of HdEG66 toward carboxymethylcellulose showed optimal temperature and pH at 38 degrees C and 6.3, respectively. cDNAs encoding HdEG66 were amplified by the polymerase chain reaction from an abalone hepatopancreas cDNA library with primers synthesized on the basis of partial amino-acid sequences of HdEG66. By overlapping the nucleotide sequences of the cDNAs, a sequence of 1898 bp in total was determined. The coding region of 1785 bp located at nucleotide position 56-1840 gave an amino-acid sequence of 594 residues including the initiation methionine. The N-terminal region of 14 residues in the deduced sequence was regarded as the signal peptide as it was absent in HdEG66 protein and showed high similarity to the consensus sequence for signal peptides of eukaryote secretory proteins. Thus, matured HdEG66 was thought to consist of 579 residues. The C-terminal region of 453 residues in HdEG66, i.e. approximately the C-terminal three quarters of the protein, showed 42-44% identity to the catalytic domains of glycoside hydrolase family 9 (GHF9)-cellulases from arthropods and Thermomonospora fusca. While the N-terminal first quarter of HdEG66 showed 27% identity to the carbohydrate-binding module (CBM) of a Cellulomonas fimi cellulase, CenA. Thus, the HdEG66 was regarded as the GHF9-cellulase possessing a family II CBM in the N-terminal region. By genomic PCR using specific primers to the 3'-terminal coding sequences of HdEG66-cDNA, a DNA of 2186 bp including three introns was amplified. This strongly suggests that the origin of HdEG66 is not from symbiotic bacteria but abalone itself.