Structures of the N-linked sugar chains in the PAS-6 glycoprotein from the bovine milk fat globule membrane

The structures of the N-linked sugar chains in the PAS-6 glycoprotein (PAS-6) from the bovine milk fat globule membrane were determined. The sugar chains were liberated from PAS-6 by hydrazinolysis, and the pyridylaminated sugar chains were separated into a neutral (6N) and two acidic chains (6M and 6D), the acidic sugar chains then being converted to neutral sugar chains (6MN and 6DN). 6N was separated into two neutral fractions (6N13 and 6N5.5), while 6MN and 6DN each gave a single fraction (6MN13 and 6DN13). The structure of 6N5.5, which was the major sugar chain in PAS-6, is proposed to be Man16 (Man13) Man14GlcNAc14GlcNAc-PA; 6N13, 6MN13 and 6DN13 are proposed to be Gal13Gal14GlcNAc12Man16 (Gal13Gal14GlcNAc12Man13) Man14GlcNAc14 (Fuc16)GlcNAc-PA;6M and 6D had 1 or 2 additional NeuAc residues at the non-reducing ends of 6MN13 and 6DN13, respectively. © 1998 Rapid Science Ltd     

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.     

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.     

Possible intrachromosomal duplication in a case of trisomy 9p

Summary A 5-year-old boy with multiple minor anomalies and mental retardation was found to have chromosomal condition of 46,XY,inv dup(9p)(pterp13::p21p24::p13qter). The clinical features of the propositus fit well with those of trisomy 9p which have been established to be a clinical entity.     

The pyridine-nucleotide cycle in tobacco

In order to elucidate the NAD-recycling pathway the following enzyme activities have been characterized in different tobacco tissues and in tomato root: NAD pyrophosphatase, nicotinamide mononucleotide (NMN)/nicotinic acid mononucleotide (NaMN) glycohydrolases, nicotinamidase and nicotinic acid phosphoribosyltransferase. The investigations were performed with protein extracts purified by gel filtration and enzymatic activities were determined by high-performance liquid chromatography methods. The kinetic parameters of the different enzymes from tobacco root and their specificity are reported. The data are in favor of the so-called pyridine-nucleotide cycle VI (NADNMNnicotinamidenicotinic acidNaMNnicotinic acid adenine dinucleotideNAD). In the nicotine-producing tobacco root a further direct route leading from NaMN to nicotinic acid is proposed. These data are reconciled with the assumption that it is nicotinic acid which is provided by the pyridine-nucleotide cycle for the synthesis of nicotine.Abbreviations HPLC high-performance liquid chromatography - Na nicotinic acid - NaAD nicotinic acid adenine dinucleotide - NaMN nicotinic acid mononucleotide - NMN nicotinamide mononucleotide - PRPP 5-phosphoribosyl-1-pyrophosphate This contribution is dedicated to Professor Augustin Betz on the occasion of his 65^th birthday     

Particle size-related phosphate binding and P-release at the sediment–water interface in a shallow German lake

Phosphate binding and P-release in the sediment of the eutrophic shallow Lake Bützow are described based on sediment profiles, particle size fractions and incubation experiments. Total phosphorus was about 15% higher in the upper 0.5 cm layer than in the 0.5–1 cm layer. Phosphorus binding varied with sediment depth. Hot P_NaOH and P_HCl were the dominant fractions in all sediment horizons down to 10 cm depth, with values ranging from 20 to 30%. The P_H2O, P_BD, o-P_NaOH and nr-P_NaOH decreased with depth. The P_BD contributed 21% to Tot-P in the horizon 0–0.5 cm and decreased by half in 1–2 cm. The greatest proportion of particles (35%) was found in the 100–200 m fraction. This size fraction also accumulated most of the phosphate. Moreover, P-forms were differently distributed in the various particle sizes of the sediment. Sediment particles <40 m can be resuspended by a wind velocity of 2 m s^–1, whereby 17% of the Tot-P from the topmost sediment were transported into the water column. The proportions of released labile phosphate, organic phosphate and hydrolysable phosphate were higher, with values of 24, 33 and 26%, respectively. Dissolved P was released under oxic and anoxic incubation, but anoxic release was higher. Comparison of the results shows that the P-release under anoxic conditions was equal to the P-release by resuspension, but under anoxic conditions the release of bioavailable P was higher.     

A Comparison of the Lipopolysaccharides and O-Specific Polysaccharides of Azospirillum brasilense Sp245 and Its Omegon-Km Mutants KM018 and KM252

The lipopolysaccharides (LPSs) extracted from the outer membrane of Azospirillum brasilense Sp245 and its Omegon-Km mutants KM018 and KM252 with a hot aqueous solution of phenol were found to differ in the content of carbohydrates, glucosamine, and total phosphorus and in the proportion of octadecenoic and hexadecanoic acids in the lipid moieties of the LPSs. The carbohydrate moieties of the LPSs were heterogeneous in charge. The analysis of the O-specific polysaccharides (O-PSs) of the mutants KM018 and KM252 by gas–liquid chromatography, IR spectroscopy, and NMR spectroscopy showed that they are composed of the same linear pentasugar repeating units 2)--D-Rhap-(1 3)--D-Rhap-(1 3)--D-Rhap-(1 2)--D-Rhap-(1 2)--D-Rhap-(1 as the O-PSs of the parent strain Sp245. The reported differences in the biological activity of the LPSs of the parent and mutant strains can be due to their different chemical composition.     

Decomposition of polyurethane in a garbage landfill leakage water and by soil microorganisms

Summary The decomposition of polyurethane, measured gravimetrially or using infrared spectrophotometry, was found to be more complete in polyurethane based on polyester and only very small in polyurethane based on polyether. In the presence of clay minerals the decomposition was inhibited. If positive, the decomposition of polyurethane followed the sequence: remaining free isocyanatesurea and amide groupsurethane groupsisocyanuric acid rings.     

Molecular characterization of β-thalassemia in Czechoslovakia

Summary We have identified different -thalassemia mutations in 93 members of 34 families of Czech or Slovakian descent using gene amplification, hybridization with specific ³²P-labeled oligonucleotide probes, sequencing of amplified DNA, and gene mapping. The GA mutation at IVS-I-1 was found in 18 families; other Mediterranean mutations were IVS-II-1 (GA), IVS-II-745 (CG), IVS-I-110 (GA), and codon 39 (CT); these were present in 9 additional families. The GT mutation at codon 121, known to cause Heinzbody -thalassemia, was present in 3 families, and the frameshift at codons 82/83 (-G), first described in the Azerbaijanian population, in 2 families. A newly discovered allele was a frameshift at codons 38/39 (-C). One -thalassemia allele was incompletely characterized. We observed in 2 families a TC mutation at position +96 UTR (untranslated region) relative to the termination codon; this mutation likely is a rare polymorphism, -Thalassemia was rare; only one person carried the -^3.7 heterozygosity, and one other had a yet to be identified -thalassemia-1, while seven had the ^{anti 3.7} triplication.     

Trans-cis isomerization of retinal and a mechanism for ion translocation in halorhodopsin

The chromophore in halorhodopsin (HR) which acts as a light-driven chloride pump in halobacteria shares many properties with its counterpart in bacteriorhodopsin (BR): (i) a similar retinal protein interaction, (ii) trans to cis isomerization and (iii) similar intermediates of its photocycle. One major difference between the two chromoproteins is that the HR chromophore does not become deprotonated during its photocycle. A mechanism for the photocycle of HR is presented, which, in close analogy to an earlier proposed mechanism for BR, involves the sequence of all-trans 13-cis, 14s-cis 13-cis all-trans isomerizations of the chromophore, a Schiff base of retinal. In contrast to the situation in BR the 13-cis, 14s-cis13-cis isomerization is induced not by deprotonation of the retinal Schiff base chromophore but rather by the movement of an anion (Cl^-) towards the protonated nitrogen of the Schiff's base. The suggested mechanism involves the Schiff base directly in the chloride translocation in halorhodopsin.     

The contribution of pigment transitions to sensitivity changes in the barnacle photoreceptor and the correlation with the prolonged depolarizing afterpotential

A conditioning light can cause a decrease (adaptation) or an increase (facilitation) in the sensitivity of barnacle photoreceptors, as measured by the amplitude of the late receptor potential (LRP). We show that a net transfer of visual pigment from the rhodopsin (R) to the metarhodopsin (M) state induces a large facilitation whereas the reverse transfer results in a much smaller facilitation or even an adaptation. These effects were not due to the response to the conditioning light but to the pigment reactions. When the conditioning light did not alter the pigment population (i.e., M M, R R) it was followed by an intermediate degree of facilitation. These conclusions are correct for cells which have relatively low sensitivity. In sensitive cells, all pigment transitions produce adaptation.LRP facilitation and the prolonged depolarizing afterpotential (PDA) show several common characteristics with respect to pigment transitions: 1.Their magnitude increases with the amount of pigment transferred from R to M. 2. Both are depressed by the M R transition. 3. Their production is impeded by the M R transition. 4. The PDA itself is facilitated by the R M transition and this facilitation decays with a time course comparable to that of LRP facilitation. These results suggest that there may be an underlying process common to LRP facilitation and PDA.     

Interstitial deletion of the short arm of chromosome 3

Summary A de novo interstitial deletion of the short arm of chromosome 3 was prenatally diagnosed in a male fetus, karyotype 46,XY,del(3)(pterp14.2::p11qter). The fetus had craniofacial dysmorphisms, a single transverse palmar crease, ulnar deviation in the wrists, cardiovascular anomalies, a slight ureteric dilatation and a mobile caecum. Our observations are compared with five other cases with interstitial deletion of the short arm of chromosome 3 to delineate further the proximal 3p deletion syndrome. The gene for beta-galactosidase-1 (GLB-1) has previously been assigned to chromosome 3(p21q21). The absence of a gene dosis effect for GLB-1 in this study indicates exclusion of GLB-1 from 3(p11p14.2).     

Leukotriene-related gene polymorphisms in ASA-intolerant asthma: an association with a haplotype of 5-lipoxygenase

A recent study has demonstrated the possible involvement of a leukotriene C4 synthase (LTC4S) gene polymorphism in ASA-intolerant asthma (AIA) in a Polish population, whereas no significant association was noted in other populations. To investigate the role of genetic polymorphism in AIA development, we screened single nucleotide polymorphisms (SNPs) of the key enzymes involved in arachidonate metabolism, and the cysteinyl leukotriene receptor 1 (CYSLTR1) in a large Korean population with AIA: 93 AIA and 181 ASA-tolerant asthma (ATA) patients, and 123 normal controls. The single-base extension method was used to genotype SNPs in 5-lipoxygenase (ALOX5, –1708GA, 21CT, 270GA, 1728GA), ALOX5-activating protein (ALOX5AP, 218AG), prostaglandin-endoperoxide synthase 2 (PTGS2, COX2, –162CG, 10TG, R228H, V511A), LTC4S (–444AC), and CYSLTR1 (927TC). Haplotype analyses were undertaken for the SNPs in ALOX5. No significant differences in allele and genotype frequencies of single SNPs were observed between the patient groups (P>0.05). However, the frequency of the ALOX5-ht1[G-C-G-A] haplotype in the AIA group was significantly higher than its frequency in the ATA group with a probability (P) of 0.01, odds ratio (OR) of 5.0, and 95% confidence interval (95%CI) of 1.54–17.9, and in the normal controls (P=0.03, OR=4.5, 95%CI=1.1–18.4), by using a dominant model. These results suggest a lack of association between the ALOX5AP, PTGS2, LTC4S, and CYSLTR1 gene polymorphisms and the AIA phenotype in the Korean population. However, the possible involvement of ALOX5-ht1[G-C-G-A] in AIA development is suggested.J.-H. Choi and H.-S. Park contributed equally to this work as first authors     

A Mathematical Model of the Cerebellar-Olivary System I: Self-Regulating Equilibrium of Climbing Fiber Activity

We use a mathematical model to investigate how climbing fiber-dependent plasticity at granule cell to Purkinje cell (grPkj) synapses in the cerebellar cortex is influenced by the synaptic organization of the cerebellar-olivary system. Based on empirical studies, grPkj synapses are assumed to decrease in strength when active during a climbing fiber input (LTD) and increase in strength when active without a climbing fiber input (LTP). Results suggest that the inhibition of climbing fibers by cerebellar output combines with LTD/P to self-regulate spontaneous climbing fiber activity to an equilibrium level at which LTP and LTD balance and the expected net change in grPkj synaptic weights is zero. The synaptic weight vector is asymptotically confined to an equilibrium hyperplane defining the set of all possible combinations of synaptic weights consistent with climbing fiber equilibrium. Results also suggest restrictions on LTP/D at grPkj synapses required to produce synaptic weights that do not drift spontaneously.     

Primary structure of hemoglobin α-chain ofColumba livia (gray wild pigeon)

Primary structure of hemoglobin of -chain ofColumba livia is presented. The separation of -chain was obtained from globin by ion-exchange chromatography (CMC-52) and reversed-phase HPLC (RP-2 column). Amino acid sequence of intact as well as tryptic digested chain was determined on gas-phase sequencer. Structure is aligned homologously with 21 other species. Among different exchanges, positions 24 (TyrLeu), 26 (AlaGly), 32 (MetLeu), 64 (AspGlu), 113 (LeuPhe), and 129 (LeuVal) are unique to pigeon hemoglobin. The various exchanges in -chain are discussed with reference to evolution and phylogeny. The results show that the order Columbiformes is evolutionarily closer to the order Anseriformes. Since the pigeon is homogeneous, having HbA (^A-chain) and lacks ^D-chain, its phylogenetic placement could be established among birds having single hemoglobin components.     

XylS domain interactions can be deduced from intraallelic dominance in double mutants of Pseudomonas putida.

Summary The XylS protein is the positive regulator of the TOL plasmid-encoded meta-cleavage pathway for the metabolism of alkylbenzoates in Pseudomonas putida. This protein is activated by a variety of benzoate analogues. To elucidate the functional domains of the regulator and their interactions, several fusions of the XylS C-terminus to MS2 polymerase and of the N-terminus to -galactosidase were constructed but all are inactive. In addition, 15 double mutant xylS genes were constructed in vitro by fusing parts of various mutant genes to produce mutant regulators exhibiting C-terminal and N-terminal amino acid substitutions. The phenotypic properties of the parental single mutant genes, and those of the double mutant genes, suggest that the C-terminal region is involved in binding to DNA sequences at the promoter of the meta-cleavage pathway operon, and that the benzoate effector binding pocket includes critical residues present at both the N-terminal and C-terminal ends of the protein. The intraallelic dominance of the Ile229 (Ser229 Ile) and Val274 (Asp274 Val) substitutions over the N-terminal His4l (Arg4l His) substitution, and the intraallelic dominance of Thr45 (Arg45 Thr) over Ile229 and Val274, support the proposal that these two regions of the regulator interact functionally. Combination of the Leu88 (Trp88 Leu) and Arg256 (Pro256 Arg) substitutions did not suppress the semiconstitutive phenotype conferred by Leu88, but resulted in a protein with altered ability to recognize benzoates. In contrast, the Leu88 semiconstitutive phenotype was suppressed by Va1288 (Asp288 Val), and the double mutant was susceptible to activation by benzoates. The results suggest that intramolecular interactions between the C- and N-terminal regions of XylS are critical for activation of the regulator by the effector.     

Specific Status of Propithecus spp.

Controversial taxonomic relationships within Propithecus have consistently made conservation and management decisions difficult. We present a multidisciplinary phylogenetic analysis of Propithecus supporting the elevation of 4 subspecies to specific status: P. diadema perrieri P. perrieri, P. diadema candidus P. candidus, P. diadema edwardsi P. edwardsi, and P. verreauxi coquereliP. coquereli; leaving P. diadema diadema as P. diadema and P. verreauxi verreauxi as P. verreauxi.     

Mutation analysis of glucose-6-phosphate dehydrogenase (G6PD) variants in Costa Rica

Summary Glucose-6-phosphate dehydrogenase (G6PD) deficiency has previously been reported among both the black and white populations of Costa Rica. All 28 G6PD A — samples were found to be of the common G6PD A-^376G/202Atype. A previously described mutation associated with nonspherocytic hemolytic anemia, G6PD Puerto Limón, was found to be due to a GA transition at nucleotide (nt) 1192, causing a glulys substitution. Mutations in this region of the G6PD molecule seem invariably to be associated with chronic hemolytic anemia. G6PD Santamaria had been described previously in two unrelated white subjects. We found that both did, indeed, have the same mutations. In this variant the AG substitution at nt 376 that is characteristic of G6PD A was present, but an AT mutation at nt 542, apparently superimposed on the ancient G6PD A mutation, resulted in an aspval substitution. Thus, the gain of a negative charge at amino acid 126 was counterbalanced by the loss of a charge at amino acid 181, giving rise to a variant with the G6PD A mutation but with normal electrophoretic mobility.     

Assignment of human phosphoribosylglycinamide synthetase locus to region 21q221

Summary The enzymatic activity of phosphoribosylglycinamide synthetase (GARS) has been studied in several cases of partial monosomies and full and partial trisomies 21. An excess of GARS activity was found in regular trisomy 21 with a trisomy 21/normal ratio equal to 1.55. A 0.99 ratio was found in 21q2121pter monosomy; a 0.54 ratio was found in 21qter21q22 monosomy; a 0.88 ratio, in 21q2121pter trisomy, and a 1.46 ratio, in 21q22.1 trisomy. Consequently, the GARS gene locus, assigned to chromosome 21, could be localized in subband 21q22.1.     

Regional mapping of catalase and Wilms tumor—aniridia,genitourinary abnormalities,and mental retardation triad loci to the chromosome segment 11p1305→p1306

Summary Gene dosage effects for catalase (CAT) were studied in two unrelated patients with an interstitial deletion involving 11p13 to determine precisely the sites of the genes for CAT and the Wilms tumor—aniridia, genitourinary abnormalities, and mental retardation triad (WAGR) in the 11p13 band. Case 1 had the aniridia-Wilms tumor association, and case 2 showed the AGR triad. The karyotypes identified by high resolution banding techniques were 46,XY,del(11)(pterp13::p11.11qter) for case 1 and 46,XY,t(2;17) (q23;q25), del(11) (pterp13::p11.2 qter) for case 2. In both cases, the distal breakpoints of the deleted chromosomes 11 appeared to have occurred on the middle portion of 11p13 (11p1305p1306). The level of erythrocyte CAT activities in case 1 was reduced (47% of normal), while that in case 2 was normal. The results suggested not only that both the CAT and WAGR should be mapped to chromosome region 11p1305p1306, but also that in this region the CAT locus is more distally placed than the WAGR locus. Because of the proximity of the two gene loci, assays of erythrocyte CAT may be useful to identify a submicroscopic deletion in some patients with sporadic aniridia and to predict a risk of developing Wilms tumor.