C.P.-D.D.-M.A.S. 13C-N.M.R. investigations of anhydrous and hydrated cyclomalto-oligosaccharides: The role of water of hydration

Hydrated and anhydrous cyclomaltohexaose, cyclomaltoheptaose, and cyclomalto-octaose (cyclodextrins) have been investigated by the c.p.-d.d.-m.a.s. ¹³C-n.m.r. technique. The chemical shifts of the signals of C-1 and C-6 provide information about conformation and the results agree fairly well with the earlier scattering data on hydrated systems, but some discrepancies have been found for cyclomaltohexaose. Th conformation of the macro-rings seems to be determined by the hydration water. The unique role of water in forming crystals of cyclomalto-oligosaccharides is demonstrated.     

1H n.m.r. studies of insulin. Assignment of resonances and properties of tyrosine residues.

The assignment of the aromatic 1H n.m.r. resonances of the four tyrosine residues of bovine 2-zinc insulin is reported, based on double resonance techniques, use of Hahn spin echo pulse sequences and examination of specific derivatives nitrated at tyrosines A14 and A19 as well as des-(B26-B30)-insulin. Titration curves of the four tyrosine residues show that residues A14 and B16 have normal pK' values of 10.3-10.6 in solution, consistent with their accessibility to solvent in monomer and dimer in the crystal. Tyrosine residues A19 and B26 have pK' values of 11.4 and exhibit other features in their titration curves that are consistent with limited accessibility to solvent and a nonpolar environment. The meta protons of residues B16 and B26 both observe the titration of a nearby tyrosine residue, probably A19. Interpretation of the n.m.r. data obtained in solution is consistent with the crystallographic data for the monomer and dimer obtained on insulin crystals [Blundell, Dodson, Hodgkin & Mercola (1972) Adv. Protein Chem. 26, 279-402].     

C.P.-M.A.S. 13C-N.M.R. study of some solid-state inclusion complexes of cyclomalto-oligosaccharides with para-substituted benzenes

Cross polarisation—magic-angle sample spinning ¹³C-n.m.r. spectral have been measured in the solid state for p-nitrophenol, p-iodophenol, and their inclusion complexes with cyclomaltohexaose, cyclomaltoheptaose, and methylated cyclomaltohexaose. Analysis of the line-shapes of the resonances and the dipolar-dephasing experiments indicate that the guest molecules undergo motion in the host cavities, whereas the host molecules are almost static. The mode and rate of guest motion depend on the size of the cavity.     

Brachyptera Group. Sp.32. C. Brachyptera. Sp.33. C.c RUFA. Sp.34. C. Troglodytes. Sp.35. C. Fulvicapilla.

A part from considerations as to where it is best wedged into the linear sequence, the brachyptera group is defined in general terms as a compact group of four small or very small species which resemble one another in many important specific characters and the other thirty-six species classified here as Cisticola in so many ways of form, coloration and behaviour as to make them best understood by classifying them also under that generic name.     

R.b.e. and o.e.r. of extended-Bragg-peak helium ions: survival and development of rat embryos.

Rats were exposed under aerobic or hypoxic conditions to 200-1200 rads of 60Co gamma-rays or extended-Bragg-peak helium ions on the eighth day of gestation. Uterine contents were examined on the twentieth day of gestation. At the 50 per cent embryonic survival level, helium ion r.b.e. was 1(.0) (aerobic) and 1(.2) (hypoxic). Maximum attainable gamma-ray and helium-ion o.e.r.s. were 2(.2) and 1(.7) respectively, indicating an oxygen-effect gain (o.e.g.) of 1(.2). At the 10 per cent survival level helium ion r.b.e. was 1(.1) (aerobic) and 1(.4) (hypoxic). Gamma-ray and helium-ion 0.e.r.s. were 2(.0) and 1(.5) respectively, indicating a helium ion o.e.g. of 1(.3). These data demonstrate that the small fraction of high-LET radiation present in this helium ion beam has a neglible effect on the aerobic r.b.e., but lowers the effective o.e.r. of the beam approximately 25 per cent relative to that of gamma-rays. Helium ions were significantly more effective than gamma-rays in killing embryos under hypoxic conditions, in producing congenital abnormalities under aerobic conditions, and in stunting foetal growth under both conditions.     

13C-n.m.r. study of C hordein.

Insoluble xylan was prepared from ground birch (Betula pubescens) pulp by alkali extraction and precipitation with ethanol. The only sugar detected after acid hydrolysis of the preparation was xylose. The insoluble xylan was used as substrate in a nephelometric assay to determine the xylanase (EC 3.2.1.8, 1,4-beta-D-xylan xylanohydrolase and EC 3.2.1.37, 1,4-beta-D-xylan xylohydrolase) activities of Aspergillus and Trichoderma enzymes. The nephelometric method is reliable in evaluating xylanase hydrolysis of insoluble xylan.     

Aspects of audit. 1. The background.

Methods of reviewing health care already exist in Britain, but the debate continues about how practical and acceptable such a review is. The many different terms used to describe review only confuse the issue. "Audit" is a useful term for describing the review of medical work by medical people. This can be divided into "internal audit," or peer review, and "external audit"--that is, review by organisations outside hospital and general practice. The concepts of internal and external audit have a great impact upon the attitudes held by the medical profession about audit. The shortcomings of audit by the professional standards review organisations in the United States are not inevitable in Britian.     

Neurobiology of Stomotoca. I. Action systems.

The layout of nerves, muscles, and conducting epithelia is described for the simple hydrozoan medusa Stomotoca. Comparisons are drawn with Sarsia and other recently studied forms. The major action systems are those responsible for swimming, crumpling (protective involution), tentacle posture, pointing (unilateral reciprocal flexions of the manubrium and margin), and visceral movements (barely mentioned). Crumpling is a simple summating response in this species. Crumpling and pointing are considered to use the same effectors but different conduction pathways. New histological results include the demonstration of a nerve plexus running through the endodermal canal system and a nerve plexus in the ectoderm encircling the peduncle. Special attention is given to the distribution of synapses and gap junctions, as possible trasmission pathways in behavioral responses. Some details are included on organization within the marginal nerve rings.     

R.M. Cowling,D.M. Richardson & S.M. Pierce (eds.) 1977. Vegetation of Southern Africa.

We determined the role of bird dispersal in seed and seedling dynamics of the tree Kalopanax pictus from 7 years of observing seed rain and seedling emergence in a broad-leaved deciduous forest in central Japan. We also performed an experiment on the influence of seed pulp on germination of seeds of K. pictus. Seeds of this species can lie dormant for several years, and this causes rather constant yearly seedling emergence in spite of irregular seed production. The spatial distribution of the seedlings that emerged each year (maximum distance from nearest conspecific seed-bearing tree of 90 m) was wider than that of gravity-dispersed seeds (max. distance of 37 m), suggesting seed dispersal by birds in winter. Emerged seedling densities at sites over 20 m from the nearest conspecific seed-bearing tree were highest in the spring of 1991, about half a year after the largest seed fall of the observation period. However, emerged seedling densities within 20 m from seed-bearing trees were highest in 1992, 1.5 years after the largest seed fall. These field observations may be explained by the experimental results on the effects of seed pulp on germination. Intact seeds germinate slowly at low germination rates, while seeds without seed pulp germinate quickly at high germination rates. Fallen seeds with seed pulp thus appear to form a seed bank near seed sources (temporal dispersal), while seeds scattered by birds appear to increase the possibility of reaching the present safe sites in distant areas with quick germination (spatial dispersal).     

Molecular analyses of the Salmonella g. . . flagellar antigen complex.

Salmonella flagellar filaments are polymers of a highly antigenic protein, termed flagellin. Eight main subfactors have been identified in the Salmonella phase-1 g. . . series flagellar antigen. To determine the molecular basis for expression of the epitopes by which the g. . . family subfactors are distinguished, 10 members of this series were selected and their fliC (the structural gene for phase-1 flagellin) genes were sequenced. Comparative analyses of the inferred primary structures of these flagellins did not allow the identification of linear epitopes responsible for the antigen subfactors. This suggests that conformational aspects are involved in determining the antigenic specificity in these cases. A phylogenetic analysis of the flagellin sequences showed that members of the g. . . series do not form a single coherent unit.     

N.m.r. and e.p.r. characterization of [4-carboxy-2,6-dinitrophenyl-lysine]cytochromes c.

Monosubstituted [4-carboxy-2,6-dinitrophenyl-lysine]cytochromes c were investigated by n.m.r. and e.p.r. Modification of Lys-13 or Lys-72 in ferricytochrome c by 4-chloro-3,5-dinitrobenzoate yields either of two different conformers that are rapidly exchanging in the native form. The equilibrium involves small local changes in the conformation of Met-80 (the sixth ligand) and Phe-82, as a result of whether Lys-13 is the 'on' or 'off' position in the Lys-13--Glu-90 salt bridge.     

Neurobiology of Polyorchis. I. Function of effector systems.

The electrical correlates of activity in the effector systems responsible for swimming, crumpling and postural changes have been recorded in the anthomedusan Polyorchis penicillatus. Motor spikes (pre-swim pulses), that initiate swimming contractions, appear without delay at distant sites on the inner nerve-ring in unstimulated preparations. Levels of Mg++ anaesthesia which block the neuromuscular junctions between PSP giant neurons and swimming muscle do not affect PSP activity. Swimming muscle potentials can be recorded from subumbrella and velar muscle sheets using extra- and intracellular electrodes. These action potentials have a distinct plateau and are propagated in a myoid fashion. Resting potentials average -70 mV with spikes overshooting zero by some 62 mV. The effects of repetitive stimulation are described. Extracellular recordings indicate that neuronal pathways may play a major role in mediating crumpling, unlike many other species where epithelial pathways are more important. Endodermal spikes recorded intracellularly from the radial and ring canals have amplitudes of some 92 mV arising from resting potentials that average -55 mV. Repetitive stimulation causes a decrease in amplitude and increase in duration of epithelial action potentials. Tentacle length is controlled by a pacemaker system located in both nerve rings. The frequency of spikes (PTPs) generated by this system determines the length and tonus of tentacles. The neuromuscular junctions between the motor neurons and tentacle muscle are Mg++ sensitive and show facilitating properties.     

Proteinases of Streptomyces fradiae. II. Specificity.

It has been shown that extracellular proteinases synthesized by a keratinolytic strain of S. fradiae are characterized by diversified activity in the decomposition of both proteins and synthetic substrates. Among the six proteinases isolated, apart from the ones dominating and having relatively low specificity, there are two enzymes characterized by narrow catalytic abilities--extremely similar to those of trypsin. These proteinases intensively degraded all the trypsin substrates studied, but they were inactive or showed slight activity toward others. They were also highly sensitive to such specific inhibitors of trypsin as TLCK, SBTI and TIO.     

On the nature of P.P.L.O.

Summary In a number of strains of the three species of P.P.L.O. of human origin coccoid elements have been observed, which are Gram positive or Gram variable and of very small size (<0.5 μ). They are able to grow in pure culture on the surface of nutrient agar in very small colonies. Although they have bacterial characteristics, they could not be identified as known bacteria. Agglutination tests are in favour of a serological relationship between the coccoid elements and P.P.L.O. The suggestion that these elements might be an outside contamination is made unlikely by several observations. The supposition that P.P.L.O.s are L forms of these coccoid elements should be considered further.     

1H-n.m.r. evaluation of the ferricytochrome c-cardiolipin interaction. Effect of superoxide radicals.

The interaction between ferricytochrome c and cardiolipin was investigated by 1H n.m.r. at 270 MHz. From the phospholipid-induced changes of the protein spectral features it is concluded that the first 2 equivalents of cardiolipin cause a conformational change at the lower part of the solvent-exposed haem edge, involving a rearrangement of the hydrogen-bond interactions of propionate 6, thus partly accounting for the lowered redox potential of cytochrome c in the presence of cardiolipin. The increased value for the pK of the alkaline isomerization of ferricytochrome c shows that cardiolipin stabilizes the native structure of the protein, indicating that the oxidized form assumes ferrocytochrome c-like properties. Peroxidation of cardiolipin by superoxide radical ions drastically decreases the protein binding to this phospholipid. The implications of this finding, and the likelihood of the ternary cytochrome c-cardiolipin-cytochrome c oxidase complex, for the binding of cytochrome c to cytochrome c oxidase in vivo, are discussed in relation to peroxidative damage following ischaemia and reperfusion.     

E.s.r. of spin-trapped radicals in aqueous solutions of peptides. Reactions of the hydroxyl radical.

The reactions of hydroxyl radicals with 30 dipeptides and several larger peptides were studied in aqueous solutions. The OH radicals were generated by U.V. photolysis of H2O2. The short-lived peptide radicals were spin-trapped using t-nitrosobutane and identified by e.s.r. For dipeptides containing the amino terminal residues glycine, alanine and phenylalanine, abstraction of the hydrogen from the carbon adjacent to the peptide nitrogen was the major process leading to the spin-adducts. Such radicals will be referred to as backbone radicals. Dipeptides with a carbonyl terminal serine residue and also glycylglutamic acid form both backbone and side-chain radicals, with the latter being formed in larger quantities. For dipeptides, side-chain radicals were detected on either the carboxyl or amino terminal residues of both. The effect of pD on the e.s.r. sectrum of the spin-adducts of glycylglycine was studied and the pK of the carboxyl group of this radical was determined to be 2.5. For (Ala)3 and (Ala)n, with an average value of n = 1800, backbone and minor side-chain radicals were observed. For ribonucleases-S-peptide, containing 20 amino acid residues, both backbone and side-chain radicals were detected.     

The precipitation of poly-L-glutamic acid. II. beta-Precipitation.

Infrared and X-ray diffraction studies have established that in the β-precipitation region of poly-L -glutamic acid the chains are in the β-conformation. Therefore, a major molecular conformational change has taken place upon precipitation. It is shown that the size of the α-helical aggregates remains constant with time in the β-region. Strong evidence can be offered to indicate that the transformation involves a transitory random-coil intermediate. Reasons are advanced, in view of the stability of the β-form, as to why two distinct precipitation regions exist.     

The genetics of self-incompatibility in Brassica campestris L. ssp. Oleifera Metzg. I. Characteristics of S-locus. Control of self-incompatibility

Self-incompatibility in Brassica campestris c.v. Arlo is controlled by a single locus sporophytic system. The identity and expression of the S alleles were determined in eight inbred and two hybrid families. It was found that co-dominance of alleles is more frequent in the stigma, whereas dominance relations between pairs of alleles predominate in the pollen. A linear order of dominance was established between six S alleles and alleles high, intermediate and low in the dominance series were recognized.In considering the variation in the expression of compatibility and the segregation ratios in inbred, F₁, F₂ and backcross progenies, the presence of a specific S allele conditioning self-fertility, or a single dominant self-compatibility factor independent of the S locus could not be established. Instead, self-compatibility in this cultivar was ascribed to the segregation of a polygenic complex which is capable of modifying the incompatibility reaction to the point of self-fertility, or to a reduction in the strength of the reaction due to the presence of S alleles low in the dominance series.     

Gelation of sickle hemoglobin. III. Nitrosyl hemoglobin.

Sickle cell nitrosyl hemoglobin was examined for gelation by an ultracentrifugal method previously described (Briehl &; Ewert, 1973) and by birefringence. In the presence of inositol hexaphosphate gelation which exhibited the endothermic temperature dependence seen in gels of deoxyhemoglobin S was observed by both techniques. In the absence of inositol hexaphosphate no gelation was observed, nor did nitrosyl hemoglobin A exhibit gelation. On the assumption that gelation is dependent on the deoxy or T (low ligand affinity) as opposed to the oxy or R (high ligand affinity) quaternary structure this supports the conclusion that nitrosyl hemoglobin S in inositol hexaphosphate assumes the T structure, in contrast to the other liganded ferrohemoglobin derivatives oxy and carbon monoxide hemoglobin. Assuming further that the quaternary structures and isomerizations are the same in hemoglobins A and S it can also be concluded that nitrosyl hemoglobin A in inositol hexaphosphate assumes the T state. Since no gelation was seen in stripped nitrosyl hemoglobin S, inositol hexaphosphate serves to effect an R to T switch in this derivative. Thus R-T isomerization in nitrosyl hemoglobin occurs without change in ligand binding at the sixth position of the heme group confirming the conclusion of Salhany (1974) and Salhany et al. (1974).Lowering of the pH toward 6 favors gelation of NO hemoglobin S as it does of deoxy and aquomethemoglobin S (Briehl &; Ewert, 1973,1974), consistent with a favoring of the T structure due to strengthening of the interchain salt bridges and the binding of inositol hexaphosphate and/or changes in site-to-site interactions on which gelation depends.     

Studies of polysaccharide fractions from the lipopolysaccharide of Pseudomonas aeruginosa N.C.T.C. 1999.

Two polymeric water-soluble fractions were isolated by gel filtration after mild acid hydrolysis of the lipopolysaccharide from Pseudomonas aeruginosa N.C.T.C. 1999. The fraction of higher molecular weight retained the O-antigenic specificity of the lipopolysaccharide and may be 'side-chain' material. This fraction was rich in N (about 10%) and gave several basic amino compounds on acid hydrolysis; fucosamine (at least 2.8% w/w) was the only specifc component identified. The fraction of lower molecular weight was a phosphorylated polysaccharide apparently corresponding to 'core' material. The major components of this fraction and their approximate molar proportions were: glucose (3-4); rhamnose (1); heptose (2); 3-deoxy-2-octulonic acid (1); galactosamine (1); alanine (1-1.5); phosphorus (6-7). In the intact lipopolysaccharide this fraction was probably linked to lipid A via a second residue of 3-deoxy-2-octulonic acid, and probably also contained additional phosphate residues and ethanolamine. The residues of 3-deoxy-2-octulonic acid were apparently substituted in the C-4 or C-5 position, and the phosphorylated heptose residues in the C-3 position. The rhamnose was mainly 2-substituted, though a little 3-substitution was detected. The glucose residues were either unsubstituted or 6-substituted. Four neutral oligosaccharides were produced by partial acid hydrolysis and were characterized by chemical, enzymic, chromatographic and mass-spectrometric methods of analysis. The structures assigned were: Glcpalpha1-6Glc; Glcpbeta1-2Rha; Rhapalpha1-6Glc; Glcpbeta1-2Rhapalpha1-6Glc. The galactosamine was substituted in the C-3 or C-4 position, the attachment of alanine was indicated, and evidence that the amino sugar linked the glucose-rhamnose region to the 'inner core' was obtained.