Reexamination of the reality or artifacts of the microtrabeculae

The polyethylene glycol (PEG) method revealed that model systems such as erythrocytes and protein solutions, which are supposed to lack structured components, exhibit lattice structures not unlike the microtrabeculae. The compactness of the lattice was dependent on the concentration of proteins. The gelated state of gelatin exhibited lattices more compact than those of the solated state at any given concentration. Comparison of images by PEG and rapid-freezing, deep-etching replica methods showed no basic differences in the ultrastructure of the intestinal epithelial cell. This indicates that the PEG method, including chemical fixation, produces little, if any, disorganization of the cytoskeleton. All of the present findings suggest that cytoplasmic protein, nonstructure-bound or structure-forming, might be present in intact cells which could form microtrabecular structures when specimens are fixed by chemical fixatives without any extractions. Therefore, the microtrabeculae should generally be regarded as a simple marker for the presence of proteinaceous macromolecules. It is also suggested that the microtrabecular lattice, as a whole, might represent a gelated state in a given compartment when another, looser lattice is simultaneously present in the same compartment, i.e., within a single cell.     

Binding of cobalamin analogs to intrinsic factor-cobalamin receptor and its prevention by R binder

It is now known that nonphysiological cobalamin analogs exist in the gastrointestinal tract, but their metabolic behavior is unclear. In this study, [57Co]cobinamide was used to study its affinity to hog intrinsic factor-cobalamin (IF-Cbl) receptor which has no species specificity against human IF-Cbl receptor, and its relation to human saliva R binder. Cobinamide was prepared from [57Co]cyanocobalamin and separated by paper chromatography. Human IF-Cbl complex was bound to IF-Cbl receptor but free cyanocobalamin was not. Although R binder-cobinamide was not bound to the IF-Cbl receptor, free cobinamide was bound to the IF-Cbl receptor to a significant extent (about one-half of IF-cyanocobalamin binding to the IF-Cbl receptor). We then investigated the binding of cobinamide to R binder and trypsin-treated R binder. Association constant of cobinamide binding to the IF-Cbl receptor was 1.0 X 10(9) M-1 which was much lower than that of cobinamide binding to trypsin-treated R binder and to untreated R binder. Further study indicated that cobinamide binding to the IF-Cbl receptor was blocked by the addition of R binder and also by trypsin-treated R binder. We conclude that one of the roles of R binder is to prevent binding of free cobalamin analogs to the IF-Cbl receptor in the gut.     

Behavior of Japanese monkey (Macaca fuscata) mothers and neonates at parturition

Parturitional behavior in 12 caged Macaca fuscatawas analyzed. Wild-caught mothers showed adequate maternal behaviors immediately following the neonate’s expulsion. Parity differences existed in the behaviors; primiparae were more idiosyncratic than were multiparae. Among multiparae, those with two or more offspring were uniformly adequate, but those with a single birth experience varied in the adequacy of the maternal care they provided at parturition. Mothers embraced and licked their neonates and had ventroventral contact with them frequently immediately after parturition but decreased these behaviors after expulsion of the placenta. In contrast, mothers showed allogrooming after consuming the placenta. Placentophagy was correlated with the level of orality represented by maternal licking behaviors. An isolation-reared primipara reacted to her newborn in a basically negative manner, although she showed little actual aggression. She showed a rapid shift in her negative behavior during the immediate postpartum period. This mother’s newborn sought contact with her, indicating the neonate’s active role in establishing a stable mother-neonate bond.     

三七中达马烷型皂甙的热不稳定性及酸水解产物

前人已证明人参和三七中富含的达马烷型人参皂甙在通常酸性水解下甙元即发生变化,而在弱酸(如50％醋酸,0.1N盐酸)条件下则形成次级皂甙。本文报道人参甙(ginseno-sides)和三七甙(notoginsenosides)的水溶液在水浴上加热亦分别形成相应的C-20位去糖基的次级皂甙。联系到人参和三七均有在蒸煮加工后C-20位去糖基皂甙收率增大的趋势,似可认为人参和三七中的这类皂甙有相当一部分是在生药的加工泡制以及提取过程中形成的次级皂甙,而不一定是植物体的原生成分。将人参甙Rb_1单体以酸水解,不仅得到主产物人参二醇(3),还分离到异去氢原人参二醇(5)、达马烷-20(22)-烯-3β,12β,26-三醇(6)、20(R)-达马烷-3β,12β,20,25-四醇(7)以及20(S)-和20(R)-原人参二醇(1、2)的混合物,从而认为这些微量成分与人参二醇一样均为达马烷型人参皂甙在酸性水解条件下C-20位糖基断裂后由真甙元的侧链转化形成的工作产物。     

The hemolymph coagulation system in invertebrate animals

A hemocyte lysate from horseshoe crab produced a gel, when exposed to Gram-negative bacterial endotoxins. This gelation reaction of the lysate, so-called Limulus test, has been widely employed as a simple and very sensitive assay method for endotoxins. Recent biochemical studies on the principle of Limulus test indicate that the hemocytes contain several serine protease zymogens, which constitute a coagulation cascade triggered by endotoxins, and that there is a (1 3)--d-glucan-mediated coagulation pathway which also results in the formation of gel. Up to now, six protein components, designated coagulogen, proclotting enzyme, factor B, factor C, factor G and anti-LPS factor, all of which are closely associated with the endotoxin-mediated coagulation pathway, have been purified and biochemically characterized. Among these components, the complete amino acid sequences of coagulogens isolated from one American and three Asian species of horseshoe crabs have been established. Moreover, the reconstitution experiment using the isolated clotting factors, C, B, proclotting enzyme and coagulogen in the presence of endotoxin, leads to the formation of coagulin get. Based on these results, we propose here a mechanism for the Limulus coagulation cascade.     

Crystallographic studies of the chicken gizzard G-actin X DNase I complex at 5A resolution

The structure of the chicken gizzard G-actin X DNase I complex has been determined at 5 A resolution by an X-ray diffraction method. Protein phases were computed by the multiple isomorphous replacement method using four heavy atom derivatives. The mean figure of merit was 0.65. Dimensions of the three molecular species, the complex, G-actin and DNase I, were determined based on the "cypress wood" models derived from the electron density map. The natures of the heavy atom binding sites are discussed in relation to the distinction between the two component molecules. The pattern of successive contacts between actin molecules observed in the present crystal seems unrelated to that found in F-actin.     

Biosynthesis of the common precursor to vasopressin and neurophysin in vitro in transplantable human oat cell carcinoma of the lung with ectopic vasopressin production

Transplantable human oat cell carcinoma cells of the lung with ectopic vasopressin production were incubated with labeled amino acids and immunoreactive neurophysins in cell extracts were analyzed by isoelectric focusing. When the cells were incubated with L-(35S)-cysteine for 20 h, one major peak (isoelectric point; pI=5.3) and several minor peaks (pI=6.1, 5.7, 5.1, 4.9 and 4.7) of labeled proteins were observed. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the relative molecular mass (Mr) of the pI 5.7 protein was estimated to be 20,000 and that of the pI 6.1 species to be 19,000, while the remainder had a Mr of approximately 10,000. The result of the pulse-labeling experiment has clearly shown that the pI 5.7 and 6.1 proteins, which have affinity for concanavalin A, are biosynthetic precursors for the smaller form of neurophysin with a pI 5.3. When subjected to limited proteolysis with trypsin, the pI 5.7 protein generated a Mr 10,000 protein and a smaller peptide. The Mr 10,000 protein thus produced was identified as neurophysin on the basis of its pH-dependent affinity for vasopressin and the migration pattern on isoelectric focusing. The smaller peptide coeluted with synthetic arginine vasopressin and bound to neurophysin suggesting that it possesses a cysteine-tyrosyl sequence at its N-terminus. Similarly, the pI 6.1 protein liberated neurophysin and vasopressin-like peptide after incubation with trypsin. These results suggests that the glycosylated protein with a pI of 5.7 and a Mr of 20,000 is the common precursor to vasopressin and neurophysin in human oat cell carcinoma of the lung with ectopic vasopressin production. The pI 6.1 protein may be an intermediate in the conversion of the precursor to vasopressin and neurophysin.     

Immunological specificity and biological action of biliary glucagon-like substance in the rabbit

A glucagon-like substance named biliary IRG2000 whose molecular weight is approximately 2,000 was isolated by gel filtration from rabbit bile. This substance showed a strong crossreactivity as equivalent to 25.7 +/- 5.1 ng/ml of porcine glucagon in RIA with antiserum 30K specificity. Biliary IRG2000 brought about a significant increase and delayed the response of blood glucose level in coexistence with porcine glucagon, though it has no appreciable effect on the glucose level when administered singly to the mouse intraperitoneally. The response with the coexistence of these materials was far greater than when porcine glucagon was given alone. In Mortimore's type rat liver perfusion, a significant rise in glucose concentration in effluent was also observed when a mixture of biliary IRG2000 and porcine glucagon was perfused. The rate of 125I-glucagon degradation was found delayed in the presence of biliary IRG2000 when examined in the rat. Thus the increase and delayed response of glucose level in coexistence of porcine glucagon with biliary IRG2000 may be explained by a suppressive effect of glucagon degradation due to biliary IRG2000.     

Long-term cultivation of amphibian melanophores. In vitro ageing and spontaneous transformation to a continuous cell line

Pure melanophore populations isolated from the tail skin of the tadpole, Rana catesbeiana, were mass cultured for a period of 2-3 years. All cell lines of amphibian melanophores studied exhibited growth crisis (in vitro ageing) followed by spontaneous transformation to a continuous cell line, as shown by changes in growth characteristics in mass culture and in clone culture, by the appearance of the cells, and by measurements of cell volumes. Even after becoming a continuous cell line, amphibian melanophores continued to have a diploid chromosome number (2n = 26) in three of four cell lines examined. The chromosome mode in one cell line, however, changed to thirty. Measurement of melanin dispersion after the addition of alpha-melanocyte-stimulating hormone suggested that the mechanism for melanin dispersion in melanophores changed during in vitro ageing.     

Phase Shift in the Potassium Uptake Rhythm of the Duckweed Lemna gibba G3 Caused by an Azide Pulse

A 6-hour application (6-hour pulse) of 1 millimolar azide significantly changed the phase of the potassium uptake rhythm of Lemna gibba G3. The phase response curve obtained was type 0 and very similar to that caused by a 6-hour pulse of low temperature (5°C) or darkness. The magnitude of the phase shift and the type of the phase response curve depended on the concentration of azide. However, 6-hour pulses of 3 millimolar cyanide or 10 micromolar (3-(3,4-dichlorophenyl)-1,1-dimethylurea) failed to shift the phase of the rhythm, while these pulses lowered the rate of carbon dioxide uptake or release. Azide, even at 3 micromolar, selectively reduced the amplitude of the rhythm without inhibiting the mean level of potassium uptake.