Isolation and1H-NMR spectroscopic identification of the glucose-containing lipid-linked precursor oligosaccharide ofN-linked carbohydrate chains

The lipid-linked precursor ofN-type glycoprotein oligosaccharides was isolated from porcine thyroid microsomes after in cubation with UDP[³H] Glucose. The carbohydrate was released from dolichol pyrophosphate by mild acid hydrolysis, purified by gel filtration and characterized by 500-MHz¹H-NMR spectroscopy in combination with enzymatic degradation. The parent oligosaccharide was found to be Glc₃Man₉Glc-NAc₂. The three glucose residues are present in the linear sequence Glcα1-2Glα1-3 Glc, the latter being α(1-3)-linked to one of the mannose residues. In order to establish the branch location of the triglucosyl unit, the parent compound was digested with jack-bean α-mannosidase. The oligosaccharide product was purified by gel filtration, and identified by¹H-NMR as Glc₃Man₅GlcNAc₂ lacking the mannose residues A, D₂, B and D₃. Therefore, the structure of the precursor oligosaccharide is as follows: $$\begin{gathered} c b a D_1 C 4 \hfill \\ Glc\alpha 1 - 2Glc\alpha 1 - 3Glc\alpha 1 - 3Man\alpha 1 - 2Man\alpha 1 - 2Man\alpha 1 \hfill \\ 3 \swarrow 3 2 1 \hfill \\ Man\alpha 1 - 2Man\alpha 1 Man\beta 1 - 4GlcNAc\beta 1 - 4GlcNAc \hfill \\ D_{2 } A 3 6 \hfill \\ Man\alpha 1 \hfill \\ 6 \hfill \\ Man\alpha 1 - 2Man\alpha 1 \nwarrow 4 \hfill \\ D_3 B \hfill \\ \end{gathered} $$      

Growth curves,morphology and ultrastructure of ten Paracoccidioides brasiliensis isolates

The yeast phase of ten P. brasiliensis isolates were studied to characterize their growth pattern, morphology and ultrastructure. Growth curves were determined after counts of total and viable fungi units (FU) during 20 days. Three growth patterns were observed: slow, reaching approximately 10–30× 10⁶ FU/tube (Pb 18, Pb 265 and PB 2); intermediate, reaching 60–150×10⁶ FU/tube (IVIC Pb 9, IVIC Pb 267, Pb SN, Pb Vitor and Pb Campo Grande) and fast, reaching 180–370×10⁶ FU/tube (Pb 2052 and Pb 192). The highest percentage of viable cells occurred on the 6th day of culture for Pb 192, Pb Campo Grande, Pb 2052 and IVIC Pb 9; on the 8th day for Pb Vitor, Pb SN, Pb 18 and IVIC Pb 267; on the 10th day for Pb 265 and on the 12th day of culture for Pb 2. Mean generation times varied from approximately 21.2 (Pb 2052) to 102.6 hours (Pb 265). The isolates showed similar morphology, except IVIC Pb 267 which did not present a typical yeast-phase at 35°C and the two fast-growing isolates (Pb 2052 and Pb 192) that presented smaller cell sizes and less tendency to clump. The ultrastructure of the isolates was similar: the cell walls presented a width of 0.1 to 0.2 °; the mitochondria presented few cristae and had equivalent patterns of distribution and morphology; the endoplasmic reticulum was scanty, presenting narrow cisternae; the vacuoles, empty or filled with electrondense material, were numerous and two to five nuclei with pores were constantly observed.     

Plasmid profiles and antibiotic susceptibility of Haemophilus pleuropneumoniae serotypes 1, 3, 5, and 7

Abstract This paper describes the plasmid profiles obtained for 73 of 96 field isolates of Haemophilus pleuropneumoniae serotypes 1, 3, 5, and 7. We also characterized the antibiotic susceptibilities of these 96 isolates. Because of the high proportion of isolates resistant to some of the antibiotics, no conclusions can be drawn as to the role of plasmids in antibiotic resistance.     

De novo t(2;13)(p24.3;q14.2) and retinoblastoma

Summary The two probes H3-8 and H2-42, known to be located in 13q14, were mapped by in situ hybridization to either side of the 13 breakpoint of an apparently balanced de novo t(2;13)(p24.3;q14.2) detected in a patient with retinoblastoma as the only phenotypic manifestation.     

Temporal mechanisms influencing gender expression and pollen flow within a self-incompatible perennial,Amianthium muscaetoxicum (Liliaceae)

Summary Temporal mechanisms that influence the synchrony of gender expression and the patterns of withinplant pollen flow were examined in Amianthium muscatoxicum. In this species self-incompatible pollinations can clog stigmas, interfere with the growth of outcrossed pollen tubes, and reduce fecundity. The majority of flowers have partial dichogamy: a two-day period of pollen dehiscence and a four-day period of pollen viability are nested in a six-day period of pistil viability. An indeterminate flowering sequence among flowers on the same plant and partial dichogamy within flowers help reduce pollen flow within the whole plant. The combined effects at both of these levels should reduce pollen wastage and lower the incidence of stigma clogging by incompatible self pollen.     

Quantity and Kinetic Properties of Ribulose 1,5-Bisphosphate Carboxylase in C3, C4, and C3-C4 Intermediate Species of Flaveria (Asteraceae)

The kinetic properties of ribulose 1,5-bisphosphate carboxylase(RuBPC) appear to have been modified during evolution of photosynthesisto adjust to changes in substrate availability. C₄ plants areconsidered to have a higher concentration of CO₂ available toRuBPC than C₃plants. In this study, the K_m(CO₂ and catalyticcapacity (k_cat) of RuBPC and the ratio of RuBPC protein to totalsoluble protein from several Flaveria species, including C₃,C₃-C₄ intermediate, and C₄ species, were determined. The C₃and intermediate species had similar K_m(CO₂) values while theC₄ species on average had higher K_m(CO₂) values. The mean ratioof K_cat/K_m for species of each group was similar, supportingthe hypothesis that changes in K_m and K_cat, are linked. Theallocation of total soluble protein to RuBPC was lowest in theC₄ Flaveria species, intermediate in the C₃-C₄ species, andhighest in the C₃ species. The results suggest that during evolutionof C₄ photosynthesis adjustments may occur in the quantity ofRuBPC prior to changes in its kinetic properties. (Received January 4, 1989; Accepted April 11, 1989)     

Viability of Cururbita pepo pollen: biophysical and structural data

During ageing of the short-lived pollen grains of Cucurbita pepo L., water loss was examined in relation to viability using biophysical (¹H-nuclear magnetic resonance, NMR) and cytological methods (fluorochromatic reaction test, freezefracture and scanning electron microscopy). A semi-logarithmic representation of the pollen weight loss demonstrated the complexity of the dehydration process. A the study of proton loss using ¹H-NMR indicated that two major releases water of had taken place, each with different flux rates. Pulse ¹H-NMR experiments showed the occurrene of non-exponential signal decay as a function of time, indicating the existence of different fractions of water in a pollen grain sample. These fractions leave the pollen grain at different times during pollen dehydration, and one of them (that of the so-called vital water) can be related to pollen viability. The quantity of protons giving a signal during pulse ¹H-NMR experiments was very low when the pollen grains were judged to be dead according to the fluorochromatic test. Freeze-fracture replicas of these dead pollen grains (less than 25% water content) showed that the plasma membrane had become detached from the intine surface; this ultrastructural feature might therefore be involved in the loss of pollen viability.Abbreviations A initial amplitude of the NMR signal - A₂ quantity of water charcterized by T_2-2 - A₅ quantity of water characterized by T_2–5 - FCR fluorochromatic reaction - NMR nuclear magnetic resonance - T₂ transverse relaxation time - T_2-2 T₂ measured with 2 ms between each pulse of radiofrequency - T_2–5 T₂ measured with 5 ms between each pulse of radiofrequency     

Inducible nitrate reductase of rice plants as a possible indicator for nitrification in water-logged paddy soils

When following the pattern of the disappearance of NH ₄ ⁺ –N from ammonium sulfate applied to the flooded soil-rice plant system (field and greenhouse experiments) during a growing season, it was observed that the lowest NH ₄ ⁺ –N level coincided with the highest value of NR activity in the leaves. Nitrate was detected in both the root and shoot systems of the rice plants and autotrophic nitrifiers (Nitrosomonas and Nitrobacter) were particularly abundant. Since it was also demonstrated in this work that the NR activity of rice plants grown with nitrate fertilization (growth chamber culture experiments) was inducible by its substrate, it can be assumed that NH ₄ ⁺ –N oxidation takes place in the water-logged soil studied. Therefore, the occurrence of the nitrification process following NH ₄ ⁺ –N fertilizer application can be predicted by thein vitro orin situ evaluation of the NR activity of the rice leaf as an indicator.     

Cytoplasm-enriched fragments ofChara: structure and electrophysiology

Summary Cytoplasm-enriched fragments prepared from internodal cells ofChara corallina by centrifugation contain membrane bound vesicles ranging in size from a few m to hundreds of m. If the fragments are incubated in artificial pond water (APW) of pH₀ above 6.5, neutral red stains the inside of many vesicles bright crimson, suggesting the presence of inward proton-pumping. In APW of pH₀ below 6 crimson vesicles are found less frequently. Under such conditions most vesicles remain unstained inside and some develop indistinct pink halos. After a few days most fragments form a central vacuole, which stains red, regardless of the pH₀. The cytoplasmic layer still contains vesicles after vacuole formation.In order to identify the membrane bounding the vesicles various fluorescent probes were applied either by injection into the fragment or directly onto the vesicles released into artificial cytoplasm. Lucifer yellow or 6 COOH-F move readily across the tonoplast in intact cells, but did not enter any vesicles. On the other hand, the fluorescent cationic stain DIOC, which is used to highlight mitochondria and especially endoplasmic reticulum, stained the vesicle membrane. Numerous elliptical or kidney shaped nuclei in the flowing cytoplasm were highlighted with DAPI. In some fragments the nuclei formed large agreggates sometimes filling the width of the fragment.Patch-clamping the vesicles in artificial cytoplasm showed the presence of several kinds of channels, some displaying similar behaviour to the K⁺ channels observed in cytoplasmic droplets.Analogous to the plasmalemma of intact cells, the fragments without vacuoles displayed electrophysiological states dominated by either K⁺ conduction, H⁺ (or OH^–) conduction or the proton pump. On the other hand, excitation transients in fragments were of low amplitude or absent altogether. Detailed comparisons of data from fragments and intact cells are shown. The effect of vacuole formation on fragment electrophysiology was also explored.     

Beyond abortion: refusal of caesarean section

Social pressures and legal restrictions are proliferating against pregnant women. A dramatic infringement on women's rights is the court ordered cesarean section, as illustrated by the case of Angela C., a terminally ill cancer patient, 26 weeks pregnant, whose refusal of a cesarean section was overridden by a District of Columbia court. The premature infant and the mother died within two days. This case epitomizes a developing judicial pattern whose ethical reasoning the author criticizes. Within the context of the right to privacy and the concept of viability, which could legally override that right, Mahowald analyzes different situations where cesarean delivery is refused. After arguing that court ordered cesarean sections are inconsistent with court refusals to force persons to undergo less invasive procedures (e.g., bone marrow donation for the benefit of family members), she proposes alternatives to the present inconsistent practice.