Isoform-Specific Membrane Translocation of Protein Kinase C After Ischemic Preconditioning

Mild cerebral anoxic/ischemic/stress insults promote tolerance and thereby protect the brain from subsequent lethal anoxic/ischemic insults. We examined whether specific activation of PKC , , , or isoforms is associated with ischemic preconditioning (IPC) in rat brain. IPC was produced by a 2-minute global cerebral ischemia. Membrane and cytosolic fractions of the hippocampi were immunoblotted using specific antibodies for PKC, , , and . PKC showed a significant translocation to the membrane fraction from 30 min to 4 h and PKC at 4 h following IPC. In contrast, the membrane/cytosol ratio of PKC showed a tendency to decrease at 30 min and 8 h, and the membrane/cytosol ratio of PKC was significantly decreased from 30 min to 24 h following IPC. These findings indicate PKC isoform-specific membrane translocations in the hippocampus after brief global brain ischemia and suggest that activation of PKC and PKC may be associated with IPC-induced tolerance in the rat hippocampus.     

Existence and uniqueness of a sharp travelling wave in degenerate non-linear diffusion Fisher-KPP equations

In this paper we use a dynamical systems approach to prove the existence of a unique critical value c ^* of the speed c for which the degenerate density-dependent diffusion equation u _ct = [D(u)u _x ] _x + g(u) has: 1. no travelling wave solutions for 0 < c < c ^*, 2. a travelling wave solution u(x, t) = (x - c ^* t) of sharp type satisfying (– ) = 1, () = 0 ^*; '(^*–) = – c ^*/D'(0), '(^*+) = 0 and 3. a continuum of travelling wave solutions of monotone decreasing front type for each c > c ^*. These fronts satisfy the boundary conditions (– ) = 1, '(– ) = (+ ) = '(+ ) = 0. We illustrate our analytical results with some numerical solutions.     

Interaction of Amyloid Beta-Protein with Anionic Phospholipids: Possible Involvement of Lys28 and C-Terminus Aliphatic Amino Acids

Fibrillar amyloid beta-protein (A) is the major protein of amyloid plaques in the brains of patients with Alzheimer's disease (AD). The mechanism by which normally produced soluble A gets fibrillized in AD is not clear. We studied the effect of neutral, zwitterionic, and anionic lipids on the fibrillization of A 1-40. We report here that acidic phospholipids such as phosphatidic acid, phosphatidylserine, phosphatidylinositol (PI), PI 4-phosphate, PI 4,5-P₂ and cardiolipin can increase the fibrillization of A, while the neutral lipids (diacylglycerol, cholesterol, cerebrosides), zwitterionic lipids (phosphatidylcholine, phosphatidylethanolamine, sphingomyelin) and anionic lipids lacking phosphate groups (sulfatides, gangliosides) do not affect A fibrillization. A was found to increase the fluorescence of 1-acyl-2-[12-[ (7-nitro-2-1, 3-benzoxadiazol-4-yl) amino] dodecanoyl]-sn-glycero-3-phosphate (NBD-PA) in a concentration-dependent manner, while no change was observed with 1-acyl-2-[12-[(7-nitro-2-1, 3-benzoxadiazol-4-yl) amino] dodecanoyl]-sn-glycero-3-phosphoethanolamine (NBD-PE). Under similar conditions, other proteins such as apolipoprotein E, gelsolin and polyglutamic acid did not interact with NBD-PA. The order of interaction of amyloid -peptides with NBD-PA was A 1-43 = A 1-42 = A 17-42 > A 1-40 = A 17-40. Other A peptides such as A 1-11, A 1-16, A 1-28, A 1-38, A 12-28, A 22-35, A 25-35, and A 31-35 did not increase the NBD-PA fluorescence. These results suggest that phosphate groups, fatty acids, and aliphatic amino acids at the C-terminus end of A 1-40/A 1-42 are essential for the interaction of A with anionic phospholipids, while hydrophilic A segment from 1-16 amino acids does not participate in this interaction. Since positively charged amino acids in A are necessary for the interaction with negatively charged phosphate groups of phospholipids, it is suggested that Lys²⁸ of A may provide anchor for the phosphate groups of lipids, while aliphatic amino acids (Val-Val-Ile-Ala) at the C-terminus of A interact with fatty acids of phospholipids.     

Das organische Fachwerk in „Kegeln“ und „Kalotten“ der Vogel-Eischale

Zusammenfassung Das zusammenhängende organische Fachwerk in Kegeln und Kalotten der Vogel-Eischale — in der Literatur als Mamillenkern beschrieben —, von uns als Loculi bezeichnet, wurde insbesondere beim Nandu (Rhea americana), daneben auch bei der Hausente (Anas domestica) an angeätzten Schalenquerschliffen mittels Goldimprägnation (s. S. 195) dargestellt. Das Fachwerk erscheint dann in den Kegeln graublau bis blauviolett, die Schalenhautfasern werden dunkelbraun oder weinrot, und die Kalotten heben sich von der umgebenden Membran in einem dunklen Mittelton ab.Jeder Loculus umhüllt in dem genannten Bereich das einzelne Calcitindividuum. Die organischen Septen des Kegelfachwerks sind im Bildungszentrum der Calcosphäriten am dicksten, weshalb dieses oft so stark imprägniert wird, daß es kaum mehr Licht durchläßt. Gegen die Säulenlage hin läuft das Kegelfachwerk allmählich aus. Die radiale Differenzierung und konzentrische Bänderung des Fachwerkes im Kegel, insbesondere sein Verhalten im Primärsphänten, werden des Näheren geschildert. In der Kalotte trennt das Fachwerk nicht nur benachbarte Calcitindividuen, sondern es grenzt diese auch gegen die Schalenhautfasern ab, welche die Kalotte durchziehen.Die Loculi entstehen aus dem organischen Anteil des schalenbildenden Sekretes, der beim Ausfallen der Calcitindividuen zurückbleibt. Beim Auswachsen drängen die Kristalle das organische Material durch einen Selbstreinigungsprozeß in die Lücken zwischen ihnen, wo es erhärtet.

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Summary The continuous loculose organic structure in cones and caps of the avian egg shell (the mamillary cores of the English literature), was thoroughly investigated in the Nandu (Rhea americana) as well as in the duck (Anas domestica) by means of etched cross-ground sections, impregnated with gold (see page 195). This procedure shows the loculi in grey-blue or violet-blue, the shell membrane fibres in brown or wine-red, and the caps in a mixed colour of both.Each loculus envelops a single calcite crystal. The greatest thickness of the organic septa is found in the crystallization centre of the calospherites; therefore the impregnation is sometimes so strong, that they have an opaque appearance. The loculi of the cones vanish gradually towards the columnar layer. The radial differentiation and the concentric striation of the loculi in the cones as well as their behaviour in the primary spherites are described in detail. In the caps the loculi separate not only neighbouring crystals, they also limite the crystals towards the shell membrane fibres, which interlace the cap.The loculi originate from the organic part of the shell-forming secretion, which remains there after the individual calcite crystals have fallen out; in a kind of self-purification process, the outgrowing crystals press the organic material into the gaps beetwen them, where its hardens.

     

Identification of a cDNA encoding a plant Lewis-type alpha1,4-fucosyltransferase

Recently, it has been found that plants, including tomato (Lycopersicon esculentum), express the Lewis-a epitope, Gal1,3(Fuc1,4)GlcNAc, on some N-glycans. By searching the EST database, it was possible to identify a tomato cDNA encoding a protein, designated FucTC, of 413 amino acids with homology to plant and mammalian 1,3/4-fucosyltransferases. The cDNA was expressed in Pichia pastoris and the recombinant enzyme was found to transfer fucose from GDP-Fuc (K_m 16 M) to lacto-N-tetraose (Gal1,3GlcNAc1,3Gal1,4Glc; K_m 80 M) as well as to 1,3- and 1,4-galactosylated N-glycans. It is concluded that FucTC is responsible for the biosynthesis of Lewis-a on N-glycans in tomato.     

Inheritance of callus formation ability in anther cultures of rice,Oryza sativa L.

Summary Inheritance of ability to form callus in rice anther culture was studied using the diallel technique. Anthers containing uninucleate microspores from two japonica cultivais (Minehikari and Taipei 309), two indica cultivars (Mingolo and Suweon 290), and 12 F₁'s of the diallel crosses involving these four parents were cultured on Chaleffs R₂ medium and evaluated for callus induction. The parents showed significant differences in anther callus formation, from 41.9% (Taipei 309) to 0% (Suweon 290). Callus induction ability was inherited as a recessive character conditioned by a single block of genes. Additive gene effects were predominant. The japonica types seemed to be good combiners for callus induction. The order of dominance among the four parents was Suweon 290, Mingolo, Minehikari and Taipei 309.     

Lactate conversion to acetate,CO2 and H2 in cell suspensions of Desulfovibrio vulgaris (Marburg): indications for the involvement of an energy driven reaction

Cell suspensions of Desulfovibrio vulgaris were found to catalyze, in the absence of sulfate, the complete conversion of 1 lactate to 1 acetate, 1 CO₂, and 2 H₂ (G₀=-8.8 kJ/mol) and of 1 pyruvate to 1 acetate, 1 CO₂, and 1 H₂ (G₀=-52 kJ/mol). Protonophores, the proton translocating ATPase inhibitor N,N-dicyclohexylcarbodiimide, and arsenate specifically inhibited H₂ formation from lactate but not from pyruvate. The results suggest that lactate oxidation to pyruvate and H₂ (G ₀=+43.2 kJ/mol) is energy driven.     

The α2 cDNA sequence of human haptoglobin carries a bacterial promoter functionalin vivo

Various constructions of human haptoglobin (Hp) cDNA coding either for the complete ^2FS precursor protein or only for the subunit have been placed under the control of the P_R promoter in the bacterial expression vector pCQV2 (Queen, 1983). In addition to the expected 45,000 dalton polypeptide synthesized after induction of the P_R promoter, the complete ^2FS constructions constitutively express a smaller polypeptide of 30,000 dalton corresponding to a truncated Hp protein. Computer analysis of the HpcDNA revealed the presence of two strong potential bacterial promoters (²P_F and ²P_S) located in the duplicated ^2FS sequence. Both Hp promoter signals are followed by potential mRNA start sites and ribosome binding sites at a compatible distance from initiation codons. In addition, the Hp² cDNA sequence, when fused upstream to the cDNA coding for ¹-antitrypsin, constitutively promotesin vivo the efficient expression of an hybrid protein specifically recognized by antibodies raised against ¹-antitrypsin or haptoglobin.     

β-Amyloid Peptide Binds Equivalently to Binary and Ternary α<Subscript>2</Subscript>-Macroglobulin–protease Complexes

₂-Macroglobulin (₂M) is a protease inhibitor that has separate binding sites for transforming growth factor- (TGF-) and -amyloid peptide (A), both of which have been identified in the ₂M sequence. In the 3D-structure of ₂M, TGF- occupies the ₂M central cavity, overlapping with the space that can accommodate up to two molecules of protease. As a result, ternary ₂M–protease complexes (2 mol protease/mol ₂M) have been reported to not bind TGF-. The goal of the present study was to test whether binding of A to ₂M is controlled by steric constraints imposed by associated proteases, similarly to TGF-. We confirmed that binary ₂M–trypsin complex (1 mol trypsin/mol ₂M) binds increased amounts of TGF-1, compared with native ₂M, while ternary ₂M–trypsin complex binds substantially decreased amounts of TGF-1. By contrast, A-binding to binary and ternary ₂M–trypsin complex was equivalent. In both cases, binding was substantially increased compared with the negligible level observed with native ₂M. Plasmin is a large protease (Mr ~82,000) that substantially occupies the ₂M central cavity; however, ₂M–plasmin complex also bound increased amounts of A, compared with native ₂M. We conclude that A accesses its binding site, in ₂M, from outside the ₂M central cavity. The TGF--and A-binding sites are spatially separated not only in the primary sequence of ₂M, but also in the 3D-structure.     

The Use of the [13C]/[12C] Ratio for the Assay of the Microbial Oxidation of Hydrocarbons

The study deals with a comparative analysis of the relative abundances of the carbon isotopes ¹²C and ¹³C in the metabolites and biomass of the Burkholderia sp. BS3702 and Pseudomonas putida BS202-p strains capable of utilizing aliphatic (n-hexadecane) and aromatic (naphthalene) hydrocarbons as sources of carbon and energy. The isotope compositions of the carbon dioxide, biomass, and exometabolites produced during the growth of Burkholderia sp. BS3702 on n-hexadecane (¹³C = –44.6 ± 0.2) were characterized by the values of ¹³C_{CO 2} = –50.2 ± 0.4, ¹³C_biom = –46.6 ± 0.4, and ¹³C_exo = –41.5 ± 0.4, respectively. The isotope compositions of the carbon dioxide, biomass, and exometabolites produced during the growth of the same bacterial strain on naphthalene (¹³C = –21 ± 0.4) were characterized by the isotope effects ¹³C_{CO 2} = –24.1 ± 0.4, ¹³C_biom = –19.2 ± 0.4, and ¹³C_exo = –19.1 ± 0.4, respectively. The possibility of using the isotope composition of metabolic carbon dioxide for the rapid monitoring of the microbial degradation of petroleum hydrocarbons in the environment is discussed.     

Average phase difference theory and 1∶1 phase entrainment in interlimb coordination

The dynamics of coupled biological oscillators can be modeled by averaging the effects of coupling over each oscillatory cycle so that the coupling depends on the phase difference between the two oscillators and not on their specific states. Average phase difference theory claims that mode locking phenomena can be predicted by the average effects of the coupling influences. As a starting point for both empirical and theoretical investigations, Rand et al. (1988) have proposed d/dt= — K sin ), with phase-locked solutions =arcsin( /K), where is the difference between the uncoupled frequencies and K is the coupling strength. Phase-locking was evaluated in three experiments using an interlimb coordination paradigm in which a person oscillates hand-held pendulums. was controlled through length differences in the left and right pendulums. The coupled frequency _c was varied by a metronome, and scaled to the eigenfrequency _v of the coupled system K was assumed to vary inversely with _c. The results indicate that: (1) and K contribute multiplicatively to (2) =0 or = regardless of K when =0; (3) 0 or regardless of when K is large (relative to ); (4) results (1) to (3) hold identically for both in phase and antiphase coordination. The results also indicate that the relevant frequency is _c/_v rather than _c. Discussion high-lighted the significance of confirming =arcsin(/K) for more general treatments of phase-locking, such as circle map dynamics, and for the 11 phase-entrainment which characterizes biological movement systems.     

Purification and properties of gammagamma-enolase from pig brain

Isoelectric focusing revealed three enolase isoforms in pig brain, which were designated as - (pI = 6.5), - (pI = 5.6), and -enolase (pI = 5.2). The pI of purified -enolase was also 5.2. The -enolase isoform of enolase was purified from pig brain by a purification protocol involving heating to 55°C for 3 min, acetone precipitation, ammonium sulfate precipitation (40%–80%), DEAE Sephadex ion-exchange chromatography (pH 6.2), and Sephadex G200 gel filtration. The final specific activity was 82 units/mg protein. As with other vertebrate enolases, -enolase from pig proved to be a dimer with a native mass of 85 kDa and a subunit mass of 45 kDa. The pH optimum for the reaction in the glycolytic direction is 7.2. The K _m values for 2-PGA, PEP, and Mg²⁺ were determined to be 0.05, 0.25, and 0.50 mM, respectively, similar to K _m values of other vertebrate enolases. The amino acid composition of pig -enolase, as determined by amino acid analysis, shows strong similarity to the compositions of -enolases from rat, human, and mouse, as determined from their amino acid sequences. Despite the differences seen with some residues, and considering the ways that the compositions were obtained, it is assumed that pig -enolase is more similar than the composition data would indicate. Moreover, it is likely that the sequences of pig -enolase and the other -enolases are almost identical. Li⁺ proved to be a noncompetitive inhibitor with either 2-PGA or Mg²⁺ as the variable substrate. This enolase crystallized in the monoclinic space group P2, or P2₁. An R _symm <5% was obtained for data between 50 and 3.65 Å, but was a disappointing 30% for data between 3.65 and 3.10 Å, indicating crystal disorder.     

ATP-dependent activation of phospholipase C by antigen,NECA, Na3VO4, and GTP-γ-S in permeabilized RBL cell ghosts: Differential augmentation by ATP,phosphoenolpyruvate and phosphocreatine

Ghosts prepared from rat basophilic leukemia cells (RBL cell ghosts) and permeabilized with -toxin fromS. aureus are a simplified system for the study of FcRI-mediated activation of phospholipase C (PLC). This activity is dependent upon ATP and magnesium, and is enhanced by the addition of another compound containing an energetic phosphate group, either phosphoenolpyruvate (PEP) or phosphocreatine (PCr). This effect appears to be specific for PEP and PCr in that other compounds with energetic phosphate bonds including fructose 1,6-bisphosphate and additional ATP are not effective. On the contrary, GTP--S, an activator of G proteins, activates PLC in the presence of ATP alone and this is not further enhanced by the addition of PEP. In addition to FcRI and GTP--S, two other stimuli lead to enhanced activity of PLC in permeabilized RBL cell ghosts: 1) an inhibitor of tyrosine phosphatases (Na₃VO₄) and 2) an analog of adenosine (NECA). Data presented here extend previous results to show that activation of PLC by GTP--S is not enhanced either by the addition of PCr or by the addition of a more MgATP. Further new findings include the observations that activation of PLC by Na₃VO₄ is augmented by PEP and PCr in a fashion similar to that observed for FcRI-mediated activation of PLC and that activation of PLC by NECA shows even more marked dependency on PEP than does activation by FcRI or Na₃VO₄. Together, these experiments demonstrate that antigen, Na₃VO₄, GTP--S, and NECA can all activate PLC in permeabilized RBL cell ghosts in the presence of ATP and that these activities are differentially affected by the addition of a second compound with an energetic phosphate bond.Abbreviations DNP₂₅BSA bovine serum albumin derivitized with 25 dinitrophenyl groups per mole of BSA - FcRI high affinity receptor for IgE - GTP--S guanosine-5-O-(3-thiotriphosphate) - IPs inositol phosphate(s) - K₂ Pipes dipotassium Pipes - Na₃VO₄ sodium vanadate - NECA 5-(N-Ethyl)carboxamidoadenosine - PCr phosphocreatine - PEP phosphoenolpyruvate - PLC phospholipase C specific for phosphoinositides - RBL rat basophilic leukemia     

Immunocytochemical analysis of phosphatidylinositol-specific phospholipase C in PC12 cells: predominance of the δ isoform during neural differentiation

The rat pheochromocytoma PC12 cell line, which differentiates into sympathetic neurons under nerve growth factor (NGF) treatment, contains at least three phosphoinositidase C (PIC) isozymes, PIC , PIC , PIC . These isozymes have been previously shown to display a different subcellular localization. To determine whether or not NGF induces changes in the presence and/or distribution of PIC isozymes during PC12 neural differentiation, studies were carried out by means of in situ immunocytochemistry. After NGF administration the proliferative activity was progressively reduced to very low levels, as measured by bromodeoxyUridine incorporation, and a neuron-like morphology was displayed by almost all cells. In unstimulated PC12 cells, PIC was detected in the nucleus whereas PIC was only cytoplasmic; PIC was found in both cell compartments. In cells treated with NGF for 3 days, neural processes extended to twice the diameter of the cell body; the isoform was concentrated near the nucleus, while the immunoreactivity of the form remained constant and the form was increased. After 10 days of treatment with NGF, PIC was hardly detectable and PIC immunostaining was considerably decreased. On the contrary, PIC progressively increased and, after 14 days of NGF exposure, fully differentiated cells displayed an intense labelling of cell body and neurites. In the same cells, PIC and PIC were almost negative. These results suggest that NGF dependent neural differentiation is related to the selective down regulation of PIC and and the increase of PIC isozyme associated with the decrease of cell proliferation.     

Cell recycling studies for α-amylase production by Bacillus amyloliquefaciens

Production of -amylase by a strain of Bacillus amyloliquefaciens was investigated in a cell recycle bioreactor incorporating a membrane filtration module for cell separation. Experimental fermentation studies with the B. amyloliquefaciens strain WA-4 clearly showed that incorporating cell recycling increased -amylase yield and volumetric productivity as compared to conventional continuous fermentation. The effect of operating conditions on -amylase production was difficult to demonstrate experimentally due to the problems of keeping the permeate and bleed rates constant over an extended period of time. Computer simulations were therefore undertaken to support the experimental data, as well as to elucidate the dynamics of -amylase production in the cell recycle bioreactor as compared to conventional chemostat and batch fermentations. Taken together, the simulations and experiments clearly showed that low bleed rate (high recycling ratio) various a high level of -amylase activity. The simulated fermentations revealed that this was especially pronounced at high recycling ratios. Volumetric productivity was maximum at a dilution rate of around 0.4 h^–1 and a high recycling ratio. The latter had to exceed 0.75 before volumetric productivity was significantly greater than with conventional chemostat fermentation.List of Symbols a proportionality constant relating the specific growth rate to the logarithm of G (h) - a ₁ reaction order with respect to starch concentration - a ₂ reaction order with respect to glucose concentration - B bleed rate (h^–1) - C starch concentration (g/l) - C ₀ starch concentration in the feed (g/l) - D dilution rate (h^–1) - D _E volumetric productivity (KNU/(mlh)) - e intracellular -amylase concentration (g/g cell mass) - E extracellular -amylase concentration (KNU/ml) - F volumetric flow rate (l/h) - G average number of genome equivalents of DNA per cell - k _l intracellular equilibrium constant - k ₂ intracellular equilibrium constant - k _s Monod saturation constant (g/l) - k ₃ excretion rate constant (h^–1) - k _d first order decay constant (h^–1) - k _gl rate constant for glucose production - k _st rate constant for starch hydrolysis - k _t1 proportionality constant for -amylase production (gmRNA/g substrate) - k ₁ translation constant (g/(g mRNAh)) - KNU kilo Novo unit - m maintenance coefficient (g substrate/(g cell massh)) - n number of binding sites for the co-repressor on the cytoplasmic repressor - Q repression function K₁/K₂Q1.0 - R ratio of recycling - R _s rate of glucose production (g/lh) - r _c rate of starch hydrolysis (g/(lh)) - R _eX retention by the filter of the compounds X: starch or -amylase - r intracellular -amylase mRNA concentration (g/g cell mass) - r _C volumetric productivity of starch (g/lh) - r _E volumetric productivity of intracellular -amylase (KNU/(g cell massh)) - r _r volumetric productivity of intracellular mRNA (g/(g cell massh)) - r _e volumetric productivity of extracellular -amylase (KNU/(mlh)) - r _s volumetric productivity of glucose (g/(lh)) - r _X volumetric productivity of cell mass (g/(lh)) - S ₀ free reducing sugar concentration in the feed (g/l) - S extracellular concentration of reducing sugar (g/1) - t time (h) - V volume (l) - X cell mass concentration (g/l) - Y yield coefficient (g cell mass/g substrate) - Y _E/S yield coefficient (KNU -amylase/g substrate) - Y _E total amount of -amylase produced (KNU) - substrate uptake (g substrate/(g cell massh)) - specific growth rate of cell mass (h^–1) - _d specific death rate of cells (h^–1) - _m maximum specific growth rate of cell mass (h^–1) This study was supported by Bioprocess Engineering Programme of the Nordic Industrial Foundation and the Center for Process Biotechnology, the Technical University of Denmark.     

A novel PH-CT-COSY methodology for measuring JPH coupling constants in unlabeled nucleic acids. Application to HIV-2 TAR RNA

A quantitative analysis of J_PH scalar couplings in nucleic acids is difficult due to small couplings to phosphorus, the extreme overlap of the sugar protons and the fast relaxation of the spins involved in the magnetization transfer. Here we present a new methodology that relies on heteronuclear Constant Time Correlation Spectroscopy (CT-COSY). The three vicinal ³J_PH3, ³J_PH5 and ³J_PH5 scalar couplings can be obtained by monitoring the intensity decay of the P_i-H3_{i – 1} peak as a function of the constant time T in a 2D correlation map. The advantage of the new method resides in the possibility of measuring the two ³J_PH5 and ³J_PH5 scalar couplings even in the presence of overlapped H5/H5 resonances, since the quantitative information is extracted from the intensity decay of the P-H3 peak. Moreover, the relaxation of the H3 proton is considerably slower than that of the H5/H5 geminal protons and the commonly populated conformations of the phosphate backbone are associated with large ³J_PH3 couplings and relatively small ³J_{PH5 / H5}. These two facts lead to optimal signal-to-noise ratio for the P-H3 correlation compared to the P-H5/H5 correlation.The heteronuclear CT-COSY experiment is suitable for oligonucleotides in the 10–15 kDa molecular mass range and has been applied to the 30mer HIV-2 TAR RNA. The methodology presented here can be used to measure P-H dipolar couplings (D_PH) as well. We will present qualitative results for the measurement of P-H_base and P-H2 dipolar couplings in the HIV-2 TAR RNA and will discuss the reasons that so far precluded the quantification of the D_PHs for the 30mer RNA.     

Structures of asparagine linked oligosaccharides of immunoglobulins (IgY) isolated from egg-yolk of Japanese quail

Structures of the Asn linked oligosaccharides of quail egg-yolk immunoglobulin (IgY) were determined in this study. Asn linked oligosaccharides were cleaved from IgY by hydrazinolysis and labelled withp-aminobenzoic acid ethyl ester (ABEE) afterN-acetylation. The ABEE labelled oligosaccharides were then fractionated by a combination of Concanavalin A-agarose column chromatography and anion exchange, normal phase and reversed phase HPLC before their structures were determined by sequential exoglycosidase digestion, methylation analysis, HPLC, and 500 MHz¹H-NMR spectroscopy. Quail IgY contained only neutral oligosaccharides of the following categories: the glucosylated oligomannose type (0.6%, Glc1-3Glc1-3Man₉GlcNAc₂; 35.6%, Glc1-3Man_7–9GlcNAc₂). oligomannose type (15.0%, with the structure Man_5–9GlcNAc₂) and biantennary complex type with core structures of-Man1-3(-Man1-6)Man1-4GlcNAc1-4GlcNAc (9.9%),-Man1-3(GlcNAc1-4)(-Man1-6)Man1-4GlcNAc1-4GlcNAc (25.1%) and-Man1-3(GlcNAc1-4)(-Man1-6)Man1-4GlcNAc1-4(Fuc1-6)GlcNAc (11.4%). Although never found in mammalian proteins, glucosylated oligosaccharides (Glc₁Man_7–9GlcNAc₂) have been located previously in hen IgY.Abbreviations IgG, IgM, IgA, IgY immunoglobulin G, M, A and Y, respectively - ABEE p-aminobenzoic acid ethyl ester     

Inheritance of the dwarf plant type in blackgram (Vigna mungo (L.) Hepper)

Summary The inheritance of the dwarf plant type was studied in blackgram (V. mungo (L.) Hepper). Type 9 has erect plant type with normal internode length. The mutant line, EMSD has reduced internode length. The F₁, F₂ and F₃ generations of a cross between Type 9 and EMSD and its reciprocal were studied. The extreme dwarf plant type appeared to be governed by a single recessive gene, dw ₁ dw ₁ with no cytoplasmic effect.Part of Ph.D. Thesis submitted by the first author     

A new mouse T-cell receptorα chain variable region family

Sequence analysis of the rearranged T-cell receptor a chain gene segments from an influenza reactive T-cell clone T2.5-5 and a hemin chloride reactive T-cell hybrid SJL-HE-1.1 have revealed a previously undescribedV gene family. We have designated this familyV 15. Southern hybridization analysis has indicated that this family most probably contains only two members, and that these are conserved in each of six mouse strains representing three previously describedV haplotypes:V ^a ,V ^b , andV ^c .     

Production and characterization of antibodies to the β-(1→6)-galactotetraosyl group and their interaction with arabinogalactan-proteins

Rabbit antisera were raised against -(16)-galactotetraose coupled to bovine serum albumin (Gal₄-BSA). The antisera reacted with arabinogalactan-proteins (AGPs) isolated from seeds, roots, or leaves of radish (Raphanus sativus L.) as revealed by immunodiffusion analysis. Extensive removal of -l-arabinofuranosyl residues from these AGPs enhanced the formation of precipitin with the antisera. The antisera did not react with such other polysaccharides as soybean arabinan-4-galactan, -(14)-galactan, and -(13)-galactan, indicating their high specificity toward the consecutive -(16)-galactosyl side chains of AGPs. The antibodies were purified by affinity chromatography on a column of immobilized -(16)-galactotetraose as ligand. The specificity of the antibodies toward consecutive (16)-linked -galactosyl residues was confirmed by enzyme-linked immunosorbent assay for hapten inhibition against Gal₄-BSA as antigen, which revealed that -(16)-galactotriose and-tetraose were potent inhibitors, while -(13)-or -(14)-galactobioses and -trioses were essentially unreactive. Electron-microscopic observation of immunogold-stained tissues demonstrated that AGPs were localized in the middle lamella as well as at the plasma membrane of primary roots of radish. Agglutination of protoplasts prepared from cotyledons occurred with the antibodies, supporting the evidence for localization of AGPs in the plasma membrane. The antibody-mediated agglutination was inhibited by addition of AGPs or -(16)-galactotetraose.Abbreviations AGP arabinogalactan-protein - BSA bovine serum albumin - ELISA enzyme-linked immunosorbent assay - FITC fluorescein isothiocyanate - Gal₃-BSA -(16)-galactotriose coupled to BSA - Gal₄-BSA -(16)-galactotetraose coupled to BSA - Ig immunoglobulin - 4-Me-GlcpA 4-O-methyl-d-glucopyranosyluronic acid - M_r relative molecular mass The authors wish to thank Dr. J. Ohnishi of Department of Biochemistry, Saitama University, for his help in preparing protoplasts.