Structural diversity of O-polysaccharides and serological classification of <Emphasis Type="Italic">Pseudomonas syringae</Emphasis> pv. <Emphasis Type="Italic">garcae</Emphasis> and other strains of genomospecies 4

Novel O-serotypes were revealed among Pseudomonas syringae pv. garcae strains by using a set of mouse monoclonal antibodies specific to the lipopolysaccharide O-polysaccharide. Structural studies showed that the O-polysaccharide of P. syringae pv. garcae NCPPB 2708 is a hitherto unknown linear L-rhamnan lacking strict regularity and having two oligosaccharide repeating units I and II, which differ in the position of substitution in one of the rhamnose residues and have the following structures: I:3)--L Rha (12)-- L Rha (12)--L-Rha-(13)--L Rha (1;II: 2)--L-Rha-(13) -L-Rha-(12)--L-Rha-(13)--L Rha (1.The branched O-polysaccharides of P. syringae pv. garcae ICMP 8047 and NCPPB 588^T have the same L-rhamnan backbone with repeating units I and II and a lateral chain of 14)- or 13)-linked residues of 3-acetamido-3,6-dideoxy-D-galactose (D-Fuc3NAc). Several monoclonal antibody epitopes associated with the L-rhamnan backbone or the lateral -D-Fuc3NAc residues were characterized.Translated from Mikrobiologiya, Vol. 73, No. 6, 2004, pp. 777–789.Original Russian Text Copyright © 2004 by Ovod, Zdorovenko, Shashkov, Kocharova, Knirel.     

Rate of free-radical oxidation of C18 diene and triene fatty acids in aqueous micellar solutions and effectiveness of beta-carotene as an inhibitor of their oxidation

The rate of accumulation of conjugated dienes of polyunsaturated fatty acids was measured during free-radical oxidation of linoleic acid (18:2n-6, LA), -linolenic acid (18:3n-3, -LNA), and -linolenic acid (18:3n-6, -LNA) initiated by 2,2"-azo-bis-(2-amidinopropane) hydrochloride in aqueous micellar solutions of sodium dodecyl sulfate and sodium cholate. It was shown that, unlike homogeneous solutions, the oxidative stability of PUFAs in aqueous dispersions increased with an increase in the extent of unsaturation. The rate of LA oxidation was more than tenfold greater than that of - and -LNA. The antioxidant activity of -carotene, in contrast to homogeneous solutions, in both micellar systems studied depended on the degree of PUFA unsaturation. We found that 5 M -carotene effectively inhibited the LA oxidation (almost by 90%), whereas the oxidation of -LNA and -LNA was not inhibited by -carotene even at much greater concentration (30 M). The paradoxical discrepancy between the extent of unsaturation and the PUFA oxidation rate, as well as a decrease in the efficiency of -carotene-dependent inhibition of oxidation of more polyunsaturated fatty acids in reactions conducted in aqueous dispersions is consistent with the model according to which the peroxyl radicals of LA and fatty acids with the doublebond number greater than two exhibit different polarity.     

The polypeptide structure and assembly of Ly-2/3 heterodimers

Mild reduction of mature, thymic Ly-2/3 heterodimers of M _r 67 000 resulted in dissociation into three individual polypeptide chains, , , and , of respective M _r values 38000, 35000, and 30000. The and chains were both immunoprecipitated by a monoclonal antibody directed to the Ly-2.1 epitope whereas the Ly-3.1 antibody bound only the chain. The possibility that the and chains of each heterodimer established their interchain links within a labile precursor protein in which a and segments were fused was considered but discounted by the finding that in mice heterozygous for both Ly-2 and Ly-3 loci, the Ly-2 product of one chromosome was not exclusively joined to Ly-3 structures coded by the same chromosome. By utilizing ionic detergents which selectively alter the charge of intrinsic membrane proteins, both Ly-2 and Ly-3 polypeptides were shown to have membrane insertion sites. It is suggested that as a consequence of their likely synthesis on membrane-bound polysomes, newly synthesized Ly-2 and Ly-3 structures accumulate within the same subcellular compartment — the membranes of the rough endoplasmic reticulum. Their elevated concentration within this space may facilitate a low affinity binding interaction between Ly-2 and Ly-3 which is later stabilized by interchain disulfide bond formation.Abbreviations used in this paper BSA bovine serum albumin - DOC sodium deoxycholate - DTT dithiothreitol - HA hemagglutinin - HTAB hexadecyltrimethylammonium bromide - RER rough endoplasmic reticulum - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis - TX100 Triton X-100     

Conformation of the circular dumbbell d〈pCGC-TT-GCG-TT〉: Structure determination and molecular dynamics

Summary The circular DNA decamer 5-dpCGC-TT-GCG-TT-3 was studied in solution by means of NMR spectroscopy and molecular dynamics in H₂O. At a temperature of 269 K, a 50/50 mixture of two dumbbell structures (denoted L2L2 and L2L4) is present. The L2L2 form contains three Watson-Crick C-G base pairs and two two-residue loops in opposite parts of the molecule. On raising the temperature from 269 K to 314 K, the L2L4 conformer becomes increasingly dominant (95% at 314 K). This conformer has a partially disrupted G(anti)-C(syn) closing base pair in the 5-GTTC-3 loop with only one remaining (solvent-accessible) hydrogen bond between NH of the cytosine dC(1) and O6 of the guanine dG(8). The opposite 5-CTTG-3 loop remains stable. The two conformers occur in slow equilibrium (rate constant 2–20 s^–1). Structure determination of the L2L2 and L2L4 forms was performed with the aid of a full relaxation matrix approach (IRMA) in combination with restrained MD. Torsional information was obtained from coupling constants. Coupling constant analysis (³J_HH, ³J_HP, ³J_CP) gave detailed information about the local geometry around backbone torsion angles , , and , revealing a relatively high flexibility of the 5-GTTC-3 loop. The values of the coupling constants are virtually temperature-independent. Weakly constrained molecular dynamics in solvent was used to sample the conformational space of the dumbbell. The relaxation matrices from the MD simulation were averaged over r^–3 to predict dynamic NOE volumes. In order to account for the 1:1 conformational mixture of L2L2 and L2L4 present at 271 K, we also included S² factors and r^–6 averaging of the r^–3-averaged relaxation matrices. On matrix averaging, the agreement of NOE volumes with experiment improved significantly for protons located in the thermodynamically less stable 5-GTTC-3 loop. The difference in stability of the 5-CTTG-3 and 5-GTTC-3 loops is mainly caused by differences in the number of potential hydrogen bonds in the minor groove and differences in stacking overlap of the base pairs closing the minihairpin loops. The syn conformation for dC(1), favored at high temperature, is stabilized by solvation in the major groove. However, the conformational properties of the dC(1) base, as deduced from R-factor analysis and MD simulations, include a large flexibility about torsion angle .     

Structure-function relationships of β- D-glucan endo- and exohydrolases from higher plants

(13),(14)--d-Glucans represent an important component of cell walls in the Poaceae family of higher plants. A number of glycoside endo- and exohydrolases is required for the depolymerization of (13),(14)--d-glucans in germinated grain or for the partial hydrolysis of the polysaccharide in elongating vegetative tissues. The enzymes include (13),(14)--d-glucan endohydrolases (EC 3.2.1.73), which are classified as family 17 glycoside hydrolases, (14)--d-glucan glucohydrolases (family 1) and -d-glucan exohydrolases (family 3). Kinetic analyses of hydrolytic reactions enable the definition of action patterns, the thermodynamics of substrate binding, and the construction of subsite maps. Mechanism-based inhibitors and substrate analogues have been used to study the spatial orientation of the substrate in the active sites of the enzymes, at the atomic level. The inhibitors and substrate analogues also allow us to define the catalytic mechanisms of the enzymes and to identify catalytic amino acid residues. Three-dimensional structures of (13),(14)--d-glucan endohydrolases, (14)--d-glucan glucohydrolases and -d-glucan exohydrolases are available or can be reliably modelled from the crystal structures of related enzymes. Substrate analogues have been diffused into crystals for solving of the three-dimensional structures of enzyme-substrate complexes. This information provides valuable insights into potential biological roles of the enzymes in the degradation of the barley (13),(14)--d-glucans during endosperm mobilization and in cell elongation.     

Darwin’s Methodological Evolution

A necessary condition for having a revolution named after you is that you are an innovator in your field. I argue that if Charles Darwin meets this condition, it is as a philosopher and methodologist. In 1991, I made the case for Darwins innovative use of thought experiment in the Origin. Here I place this innovative practice in the context of Darwins methodological commitments, trace its origins back into Darwins notebooks, and pursue Darwins suggestion that it owes its inspiration to Charles Lyell.     

Histochemical properties and innervation pattern of fast and slow-tonic fibre types of the anterior latissimus dorsi muscle of the chick

Summary The anterior latissimus dorsi muscle of the chick is largely composed of slow-tonic fibres but contains a few fast fibres defined by their acid-labile, alkali-stable myofibrillar ATPase activity. These fibres are referred to as fibres. Differing from the slow-tonic ( and ) fibres which are multiply innervated withen grape endings, the (fast) fibres are innervated by typicalen plaque endings. The innervation of thirty-two fibres were studied closely and it was concluded that about a half of the fibres were focally innervated whereas a half were innervated in two distinct loci byen plaque endings. In only one case, a fast fibre with three widely spaceden plaque endings was observed. The mean intersynaptic length was significantly larger on fibres than on and fibres. No variation in the histochemical properties of myofibrillar ATPase was observed either along the entire length of singly and dually innervated fibres, or along and fibres. It is concluded that the three extrafusal fibre types of the anterior latissimus dorsi muscle maintain uniform histochemical characteristics along their length in spite of a possible innervation by several motoneurons.     

The taxonomic significance of the trunk limbs of the chydoridae (Cladocera)

Summary The differences in the structure of the trunk limbs allow to outline three sections of Chydoridae (see table I and fig. 1), coinciding with the sections distinguished according to the structure of the head pores.

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Chydoridae (Cladocera)

Chydoridae (. ), , .

     

Kinetic resolution of N-protected amino acid esters in organic solvents catalyzed by a stable industrial alkaline protease

Summary An industrial alkaline protease Alcalase has been found to be very stable in organic solvents and usable as a catalyst for resolution of N-protected amino acids, in both aqueous solution and organic solvent with high yield and optical purity. Only the L-amino acid ester has been hydrolysed.Abbreviation Cbz- carbobenzyloxy- - OMe methyl ester - Hop homophenylalanine - Nol norleucine - Aba -amino butyric acid - Nov norvaline - Fug furylglycine     

A note on dimensionless life histories for birds versus mammals

Summary Offspring production over the adult lifespan (b/M whereb is the yearly fledgling or offspring production and 1/M is the mean adult lifespan) is an approximate invariant within both birds and mammals. The two taxa differ, however, in that mammals have bothM and b as invariants (b/M = b/M) while birds do not ( is the age at first breeding). Birds have a surprising cancellation in that bothM andb are ^–0.25.     

Effects of exogenous linoleic acid on fatty acid composition,receptor-mediated cAMP formation,and transport functions in rat astrocytes in primary culture

We have examined the effects of culturing neonatal rat-brain astrocytes in medium containing delipidated serum, with or without added linoleic acid (LA, 18:26), on membrane fatty-acid composition and functions. After 18–21 days in culture, polyunsaturated fatty acids (PUFA) constituted24 mol% of the total fatty acids in the astrocytes grown in delipidated media (controls); these proportions were increased by 35–40% to33 mol% when the cells were supplemented with 35M LA. Notable differences in the PUFA profiles of the cells cultured with or without added LA included: (a) higher proportions of 6 PUFA in the LA-supplemented astrocytes (25%, relative to10% in controls) that were accompanied by an increase in the ratio of 6/3 PUFA (from <2 in controls to 5), and (b) higher proportions of 20:39 and 22:39 in the control astrocytes (>5%) relative to the LA-supplemented cells (1%). The major metabolites in the 6 PUFA-enriched cells were arachidonic (20:46), adrenic (22:46) and docosapentaenoic (22:56) acids (15, 5 & 3 mol%, respectively). Enrichment of the astrocytes in 6 PUFA did not alter basal levels of cAMP, nor did it affect the amounts of cAMP formed in response to forskolin, isoproterenol, adenosine or histamine. However, dopamine-dependent increases in cAMP formation in the presence of the phosphodiesterase inhibitor, Ro 20-1724, were reduced by 25% relative to those in controls. LA supplementation modified uptake of [³H]adenosine into the astrocytes; values for K_t for a high affinity transport were increased relative to controls, and maximum capacity of a lower affinity process was reduced. Uptake of [³H]glutamate was not altered in the 6 PUFA-enriched astrocytes. This study demonstrated that cultured astrocytes take up exogenous linoleic acid and incorporate its metabolites into, phospholipid, and that the resulting changes in membrans PUFA composition modify only specific cell functional properties.Abbreviations PUFA polyunsaturated fatty acid(s) - EFA essential fatty acid(s) - LA linoleic acid - AA arachidonic acid - DHA docosahexaenoic acid - BSA bovine serum albumin - DMEM Dulbecco's modified Eagle's medium - TBARS thiobarbituric-acid-reactive substances - NECA 5-N-ethylcarboxamidoadenosine Special issue dedicated to Dr. Leon S. Wolfe.     

Isolation and partial sequence of goat spleen prothymosin α

Goat prothymosin , a highly acidic polypeptide of pl 3.5, 109 amino acid residues, has been isolated from lymphoid and non-lymphoid tissues of young female goats. Unlike rat, murine and porcine prothymosins , goat prothymosin appears at a higher concentration in the spleen compared with the thymus. The sequence of segments of the polypeptide involving known mutations has been determined, by automatic sequencing of its tryptic peptide fragments. The acidic amino acid-rich segment in the middle of the molecule, including residues 49–83, has not been sequenced. Goat prothymosin closely resembles bovine prothymosin , with only one substitution, proline for alanine at position 85. It also resembles human prothymosin , with only three substitutions. It differs more significantly from rat and murine prothymosins , by two deletions and three substitutions. The results show the highly conserved nature of the molecule, with substitutions at given positions only.Abbreviations ProT Prothymosin - T₁ Thymosin ₁ - MLR Mixed Lymphocyte Response - HPLC High Performance Liquid Chromatography - RIA Radioimmunoassay - B Aspartic acid or Asparagine - Z Glutamic acid or Glutamine     

Moraxellae: Identification

According to published reports, all strains ofMoraxella osloensis and Moraxella (Oligella) urethralis grew on a mineral-base medium supplemented with 0.015M (0.123%) sodium acetate, but not all alkalinized a mineral-base medium supplemented with 0.008% bromthymol blue and 0.2% sodium acetate. This seeming discrepancy was examined. The growth of most strains ofM. urethralis was inhibited by 0.008% bromthymol blue, and some by 0.2% anhydrous sodium acetate. All (42) strains ofM. osloensis andM. urethralis alkalinized a mineral-base medium supplemented with 0.1% sodium acetate trihydrate and 0.001% bromthymol blue or phenol red.     

Limits of artificial selection under unbalanced mating systems

Summary The influence of unbalanced mating systems — factorial mating (FM) and random loss of families after a full diallel crossing (RS) — on the ultimate probability of gene fixation (u()) and the time required to fix or lose a gene (t()) are investigated. The average u() of these systems is smaller than that of random mating, and the range of u() for a given initial parental genotype combination is very large (close to one for most initial genotypic combinations). The average u() of different parental genotypic combinations of a given gene frequency are different. These systems accelerate the t().This article was written and prepared by U.S. Government employees on official time, and it is therefore in the public domain     

Electrochemical potential of protons in vesicles reconstituted from purified,proton-translocating adenosine triphosphatase

Summary Measurements were made of the difference in the electrochemical potential of protons ( ) across the membrane of vesicles reconstituted from the ATPase complex (TF ₀ ·F ₁) purified from a thermophilic bacterium and P-lipids. Two fluorescent dyes, anilinonaphthalene sulfonate (ANS) and 9-aminoacridine (9AA) were used as probes for measuring the membrane potential () and pH difference across the membrane ( pH), respectively.In the presence of Tris buffer the maximal and no pH were produced, while in the presence of the permeant anion NO ₃ ^– the maximal pH and a low were produced by the addition of ATP. When the ATP concentration was 0.24mm, the was 140–150 mV (positive inside) in Tris buffer, and the pH was 2.9–3.5 units (acidic inside) in the presence of NO ₃ ^– . Addition of a saturating amount of ATP produced somewhat larger and pH values, and the attained was about 310 mV.By trapping pH indicators in the vesicles during their reconstitution it was found that the pH inside the vesicles was pH 4–5 during ATP hydrolysis.The effects of energy transfer inhibitors, uncouplers, ionophores, and permeant anions on these vesicles were studied.     

Genetic analysis of brown planthopper resistance in twenty varieties of rice,Oryza saliva L.

Summary The inheritance of resistance to brown planthopper, Nilaparvata lugens (Stol.), of 20 rice cultivars was studied. Single dominant genes that are allelic to Bph 3 condition the resistance in cultivars Ptb 19, Gangala (Acc. 7733), Gangala (Acc. 15207), Horana Mawee, Kuruhondarwala, Mudu Kiriyal and Muthumanikam. Single recessive genes that are allelic to bph 4 govern the resistance in cultivars Gambada Samba, Heenhoranamawee, Hotel Samba, Kahata Samba, Kalukuruwee, Lekam Samba, Senawee, Sulai, Thirissa and Vellai Illankali. The resistance in Ptb 33, Sudu Hondarwala, and Sinna Sivappu is governed by one dominant and one recessive gene which segregate independently of each other. The dominant resistance genes in these cultivars appear allelic to either Bph 1 or Bph 3. Similarly, the recessive genes in these cultivars seem allelic to either bph 2 or bph 4. Further investigations are needed to conclusively determine the allelic relationships of resistance genes in Ptb 33, Sudu Hondarwala and Sinna Sivappu.     

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.     

On evolution of the photosynthetic pigments

Some considerations on the origin of the photosynthetic pigment apparatus in the period of transition of organisms from heterotrophic to phototrophic mode of life and further evolution of this apparatus are given in this paper.Some ideas about interrelation between major and auxiliary pigments and about different forms of the pigments in vivo are discussed from an evolutionary point of view.Special Symposium on Photochemistry and the Origins of Life, Sixth International Congress on Photobiology, Bochum, Germany.     

Sodium channel opening as a precursor to inactivation

The time constant of the process producing the delay in Na inactivation development as determined by the two pulse method (_delay) was extracted and compared to that of the slowest Na activation process ₃ for the I _Na during the conditioning pulse of that same determination. _delay and two pulse inactivation _c values were computer generated using a nonlinear least squares algorithm. _h and single pulse inactivation _h values were independently generated for each determination also with the aid of the computer using the same non-linear least squares algorithm. In one determination at 2 mV, _c was 4.68 and _delay 0.494 ms while _h was 4.70 and ₃ 0.491 ms for a _c/_h of 0.996 and a _delay/₃ of 1.006. Mean _delay/₃ from five determinations in four axons, both Cs and K perfused, and spanning a potential range of-27 to 2mV was 1.068. The precursor process to inactivation is channel opening. Some fraction of channels presumably inactivate via another route where prior channel opening is not required.     

Two-way selection for mating activity in the Mediterranean fruit fly, Ceratitis capitata

No significant variation in mating activity was observed among eight laboratory strains of the Mediterranean fruit fly (medfly), Ceratitis capitata (Wiedemann) (Diptera: Tephritidae). Two methods were used to select strains showing high and low mating activity: a single pair technique (SP) assessing time to mating and a mass technique (M) based on a mating index. Reciprocal pairings between fast and slow selected lines showed that the difference between the SP lines was female-determined whereas the difference between the M lines depended on the behaviour of both sexes. M selection, irrespective of its direction, affected two courtship parameters, vibration distance and vibration duration both of which tended to be shorter. M selection was also associated with a reduction in startle activity in females.

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Résumé Aucune variation significative n'a été observée dans les activités sexuelles de huit souches de laboratoire de la mouche méditerranéenne du fruit, Ceratitis capitata (Wiedemann) (Diptera Tephritidae).Deux méthodes ont été utilisées pour sélectionner des souches manifestant une activité sexuelle basse ou élevée: une technique utilisant un seul couple (SP), basée sur la durée de la période avant copulation, et une technique de masse (M), basée sur un index d'accouplement.Des appariements réciproques entre les lignées rapides et lentes ont montré que la différence entre les lignées SP était déterminée par les femelles, tandis que la différence entre les lignées M résultait du comportement des deux sexes. La sélection M, quelle que soit sa direction, affects deux paramètres, la distance entre partenaires lors de la vibration et la durée de la vibration qui tous deux tendent à se raccourcir. La sélection M est aussi associée à une réduction de la réaction des femelles à un brusque stimulus lumineux (Startle activity).