Mammalian-lung galactan

Exopolysaccharides of Agrobacterium tumefaciens and Rhizobium meliloti, containing d-glucose, d-galactose, pyruvic acid, and O-acetyl groups in the approximate proportions 6:1:1:1.5, were analysed by methylation. They were found to contain the following main structural units (all β-glycosidic): chain residues of (1→3)-linked d-glucose (24%), (1→3)-linked d-galactose (15%), (1→4)-linked d-glucose (20%), and (1→6)-linked d-glucose (18%); (1→4,1→6)-linked branching residues of d-glucose (12%), and terminal d-glucose residues substituted at positions 4 and 6 by pyruvate (11%). Uronic acid-containing exopolysaccharides of Rhizobium leguminosarum, R. phaseoli, and R. trifolii contained d-glucose, d-glucuronic acid, d-galactose, pyruvic acid, and O-acetyl groups in the approximate proportions 5:2:1:2:3. Methylation gave identical patterns of methylated sugar components, from which the following structural elements were deduced: chain residues of (1→3)-linked d-glucose substituted at positions 4 and 6 by pyruvate (13%), (1→4)-linked d-glucose (32%), and (1→4)-linked d-glucuronic acid (20%); (1→4,1→6)-linked branching residues of d-galactose and/or d-glucose (13%), and terminal d-glucose and/or d-galactose residues substituted at positions 4 and 6 by pyruvate (13%).     

The utilization of bathocuproinedisulfonic acid as a reagent for determining D-glucose and D-galactose levels in glycoconjugates

A sensitive, rapid, and reliable method for measuring d-glucose and d-galactose levels in glycoconjugates has been developed. In this method, the NAD(P)H produced from the enzymatic oxidation of the monosaccharides is reacted with a CuSO₄-bathocuproinedisulfonic acid reagent (Cu-BCS) to produce a color complex absorbing maximally at 486 nm. With galactose dehydrogenase and glucose dehydrogenase serving as the model enzymes, graphs of absorbance versus varying d-glucose or d-galactose concentrations yielded a linear plot from 2.5 to 250 nmol of sugar. Using this procedure, sugar released by acid hydrolysis from lactose, porcine submaxillary mucin and raffinose was quantified. When p-nitrophenyl-α-d-glucopyranoside and p-nitrophenyl-β-d-galactopyranoside were acid hydrolyzed and assayed with the Cu-BCS reagent, the amount of sugar released from each of the p-nitrophenyl compounds was found to be equal to the levels of p-nitrophenol in solution. This method is easy to use and with minor modifications can be employed for the quantification of d-glucose and d-galactose in other glycoconjugates.     

Polysaccharide components of the seed-coat mucilage from hyptis suaveolens

The mucilage isolated from the seed coat of Hyptis suaveolens contains l-fucose, d-xylose, d-mannose, d-galactose, d-glucose and 4-O-methyl-d-glucuronic acid in the mol ratios 1.0:2.5:1.5:7.0:12.5:1.1. Fractionation of the mucilage with Fehling's solution gave a neutral and an acidic polysaccharide. The neutral polysaccharide appears to be homogeneous and is composed of d-mannose, d-galactose and d-glucose in the mol ratios 1.0:4.5:7.5. The acidic polysaccharide is composed of l-fucose, d-xylose and 4-O-methyl-d-glucuronic acid in the mol ratios 1.0:2.5:1.1. It is homogeneous on gel filtration, DEAE-cellulose chromatography, sedimentation analysis and electrophoresis.     

Isolation of a collagen fraction from the body-wall glycoproteins of the leech (Hirudo medicinalis), and characterization of its carbohydrate-amino acid portion

Fractionation of the leech (Hirudo medicinalis) body-wall glycoproteins yielded a collagen fraction containing only d-glucose and d-galactose as its carbohydrate constituents. Digestion of the collagen with trypsin and pronase, and alkaline degradation of the resulting glycopeptides, gave a product that contained a disaccharide linked to hydroxylysine. Mild, acid hydrolysis of the N-acetylated glycopeptides yielded a disaccharide consisting of a d-glucose and a d-galactose residue. Various chemical and enzymic reactions of the disaccharide, the glycosyloxylysine, and the glycopeptide fraction indicated that the disaccharide is 2-O-α-d-glucopyranosyl-d-galactose, and that this is β-glycosidically linked to O-5 of the hydroxylysine residue in the collagen.     

Characterization of 5-aminolevulinate synthase from Agrobacterium radiobacter,screening new inhibitors for 5-aminolevulinate dehydratase from Escherichia coli and their potential use for high 5-aminolevulinate production

The hemA gene encoding 5-aminolevulinate synthase (ALAS) from Agrobacterium radiobacter zju-0121 showed 92.6% homology with that from A. radiobacter ATCC4718 and contained several rare codons. To enhance the expression of this gene, Escherichia coli Rosetta(DE3), which is a rare codon optimizer strain, was used as the host to construct an efficient recombinant strain. And the encoded protein was over-expressed as fusion protein and was purified by affinity purification on Ni-NTA agarose and by gel filtration chromatography on Sephadex G-25 Medium resin. The recombinant protein was partly characterized, and d-glucose, d-fructose, d-xylose, d-mannose, l-arabinose, d-galactose, lactose, sucrose and maltose were detected to have no distinct inhibition on this recombinant ALAS. Meanwhile, 20 mM d-glucose or d-xylose inhibited about 20% activity of ALA dehydratase (ALAD) from Escherichia coli Rosetta(DE3). Combining d-xylose as a new inhibitor for ALAD with d-glucose in fed-batch culture and based on the optimal culture system using Rosetta(DE3)/pET28a-hemA, the yield of ALA achieved was 7.3 g/l (56 mM) under the appropriate conditions in the fermenter.     

The effect of the strongly bound protein fraction on sugar transport in human erythrocyte ghosts

The protein fraction released from human erythrocyte membranes with 90% acetic acid enhanced the transport of several sugar species when enclosed in erythrocyte ghosts. Both the influx and the efflux of d-glucose were increased so that permeation rather than sugar binding was involved. The permeation increase was selective, being found with d-glucose, d-galactose and d-xylose but not with l-glucose or lactose. The protein-dependent sugar transport was saturable and the incorporation of proteins into the ghost membrane brought J_max to the level corresponding to intact erythrocytes, leaving K_m unchanged.     

Enzymatic syntheses of dopamine glycosides

Glucoamylase from Rhizopus mold and β-glucosidase catalysed synthesis of dopamine-d-glucoside was optimized in terms of pH, buffer, enzyme, dopamine concentrations and incubation period in organic media. Under the optimum conditions, β-glucosidase showed regioselectivity by giving rise to 3-hydroxy-4-O-(β-d-glucopyranosyl)phenylethylamine 5b. Presence of hydroxyl group at 3rd and 4th position of phenyl ring showed only monoglycosylation/arylation products with d-glucose 2, d-galactose 3 and d-mannose 4 for both the enzymes. Synthesized glycosides also exhibited ACE inhibition and antioxidant activities.     

Ethanol production from d-galactose and glycerol by Pachysolen tannophilus

Pachysolen tannophilus has recently been shown to be able to convert d-xylose, a pentose, to ethanol. Previously, d-xylose had been considered to be nonfermentable by yeasts. The present study shows that the organism can be used to obtain ethanol from other carbohydrates previously considered as nonfermentable, either by P. tannophilus in particular, d-galactose, or by yeasts in general, glycerol. Such identification for d-galactose allows P. tannophilus to be considered for fermentation of four of the five major plant monosaccharides: d-glucose, d-mannose, d-galactose and d-xylose. The ability to ferment glycerol is of potential use, in part, for the conversion of glycerol derived from algae into ethanol.     

Characterization of O-trimethylsilyl derivatives of d-glucose,d-galactose,and d-mannose by gas-liquid chromatography-chemical-ionization mass spectrometry with ammonia as reagent gas

The per(trimethylsilyl) ethers of d-glucose, d-galactose, and d-mannose were analyzed by g.l.c.-c.i.m.s. with ammonia as the reagent gas. C.i.m.s. gave simple fragmentation and fragment ions of high intensity in the high-mass range where the QM⁺ ion is also detected. The β-d anomers gave ions at m/e 558 showing intensities 3–12 times those of the α-d anomers. The epimers could be distinguished by differences in the intensities of the ions and by the observation that d-glucose gave a base peak at m/e 198, d-galactose at m/e 468, and d-mannose at m/e 204. The pyranose and furanose structures could be distinguished by comparing the ion intensities at m/e 198, m/e 271, m/e 361, m/e 396, and m/e 451. A similar analysis was also performed with 2-methylpropane as the reagent gas.     

C-(polyacetoxy)alkyloxadiazolines and related compounds

The (phenylacetyl)hydrazones of d-galactose, d-glucose, d-mannose, d-arabinose, l-arabinose, d-xylose, and l-sorbose were prepared. The d-galactose and d-arabinose derivatives were converted into their per-O-acetylated derivatives (8 and 9, respectively). The acyclic structure of 8 was proved from its direct preparation by the condensation of(phenylacetyl)hydrazine with penta-O-acetyl-aldehydo-d-galactose. Cyclization of 2,3,4,5,6-penta-O-acetyl-aldehydo-d-galactose (phenylacetyl)hydrazone with boiling acetic anhydride yielded a mixture of two products that could be separated by fractional recrystallization, to give 3-acetyl-5-benzyl-2-(polyacetoxy)alkyl-1,3,4-oxadiazolines; a mechanism for the reaction was proposed. The n.m.r. and mass spectra of some of these derivatives were discussed.     

Specificity of d-glucose transport by the apical membrane of Nereis diversicolor epidermis

(1) The specificity of d-[6-³H]glucose influx by a Na⁺-dependent and phlorizin-sensitive transport system in the apical epidermal membrane of the polychaete worm, Nereis diversicolor, was investigated in vivo. (2) The inhibitory effect of eleven d-glucose analogues on d-[6-³H]glucose influx from a 5 μM external concentration was recorded. The inhibitors (each tested at 5, 50, 500 and 5000 μM) were selected to illuminate the configurational requirements for interaction with the d-glucose transport system. (3) The following compounds were found to be significant inhibitors: methyl α-d-glucoside, methyl β-d-glucoside, d-galactose, $\text{3-O-}\text{methyl-}\text{d}\text{-glucose}$, 2-deoxy-d-glucose, d-xylose, myo-inositol, β-d-fructose; the effect was graded according to inhibitor concentration. l-Glucose also inhibited d-glucose influx but to the same extent at all four concentrations tested, suggesting transport site heterogeneity. d-Mannose and l-arabinose did not inhibit influx. (4) The most potent inhibitor, methyl-α-d-glucoside, was itself a substrate, and its transport was inhibited by phlorizin and d-glucose, as well as by substitution of Na⁺ in the incubation medium with Li⁺ or choline⁺. (5) We conclude that the specificity of the Na⁺-dependent d-glucose transporter in the apical epidermal membrane of Nereis is similar to that in the apical membrane of vertebrate small intestinal and proximal tubular epithelium, and in the tapeworm integument.     

Synthesis and growth regulator activity of fatty acyl derivatives of d-glucose and d-galactose

In an attempt to gain information about one or more components of the brassin complex, fatty acid esters of d-glucose and d-galactose were prepared and tested for growth regulator activity in a bean hypocotyl bioassay. 4-O-Acyl-d-glucoses and, perhaps, 1-O-acyl- d-galactoses had a similar qualitative activity to that of the brassin complex. 3-O-Acyl- d-galactoses inhibited elongation of bean hypocotyls and stimulated rooting. 3- And 6-O- acyl-d-glucoses both stimulated and inhibited elongation, depending on the source of fatty acids; in both cases, stimulation was observed when safflower oil was used as the source of fatty acids and inhibition was observed when peanut oil was used as the source of fatty acids. Fatty alkyl β-d-galactopyranosides were inactive.     

Structural studies on Klebsiella type 61 capsular polysaccharide

The capsular polysaccharide from klebsiella type 61 was found to contain d-galactose, d-glucose, d-mannose, and d-glucuronic acid in the ratios 1:2:1:1. Acid hydrolysis of the polysaccharide gave one aldobiouronic acid, whose structure was established. Methylation analysis of the polysaccharide provided information about the linkages in the polysaccharide. The polysaccharide is composed of a pentasaccharide repeating unit for which structures are proposed.     

Structure and biodegradation of the polysaccharide components of egg masses isolated from snails of an Ampullarius species

A polysaccharide preparation, isolated from egg masses deposited by snails of an Ampullarius species, was purified via precipitation with Cetavlon in the presence of sodium borate, and found to contain d-galactose and a smaller proportion of d-glucose, and to have two components with sedimentation coefficients of 10S and 40S. A polysaccharide, isolated from freshly laid egg masses, was highly branched and was shown to contain nonreducing α-d-glucopyranosyl and β-d-galactopyranosyl end-groups, and 3,6-di-O-substituted β-d-galactopyranosyl residues. One or more of the polysaccharide components was a d-glucopyrano-d-galactopyranan with non-reducing α-d-glucopyranosyl end-groups (1→4)-linked to β-d-galactopyranosyl residues. The polysaccharide preparations, obtained from freshly laid egg masses and from those that were left for 10 and 15 days after being laid, were structurally different from each other. With the passage of time, progressive diminution of the 10S component and the proportion of d-glucose in the polysaccharide took place, suggesting that each constituent was consumed preferentially by the snail embryos as an energy source.     

Biogenesis of the C-Glycoside leucodrin in Leucadendron Argenteum

Biogenetic pathways for leucodrin formation in Leucadendron spp. are discussed and were tested by feeding labelled d-galactose, d-glucose and l-ascorbic acid to cut stem ends of L. argenteum. It was shown that the two aldohexoses are better precursors and preferentially incorporated into that portion of leucodrin derivable from p-coumaric acid. When this route is suppressed by feeding the labelled sugars in the presence of a large amount of phenylalanine) label is predominantly incorporated in the carbohydrate part of leucodrin. The biogenesis of leucodrin therefore may proceed via coupling of l-galactono-γ-lactone and p-coumaric acid.     

Some properties of polysaccharides of microorganisms from degraded straw

Extracellular polysachcarides from bacteria and yeasts isolated from decomposed straw contained various proportions of d-galactose, d-glucose, d-mannose, uronic acid, d-xylose, l-fucose and l-rhamnose. Molecular weights of the polymers determined by viscometry and gel filtration were in the range 40 000–1800 000. All the polysaccharides stabilized aggregates of volcanic ash and most were more effective than the polysaccharide from Lipomyces starkeyi. Effectiveness seemed to be more related to molecular weight than to chemical composition.     

Carbohydrates of Padina tetrastromatica

Extraction with hydrochloric acid (pH 2.5) of the brown alga Padina tetrastromatica afforded water-soluble and water-insoluble polysaccharides. The water-soluble polysaccharide was fractionated using cetyltritmethyl ammonium bromide and chromatography on DEAE-cellulose and Sephadex G-100. A neutral laminaran like glucan and two new sulphated heteropolysaccharides comprising d-glucuronic acid, l-fucose, l-rhamnose, d-xylose, d-arabinose, d-galactose, d-glucose and half-ester sulphate were obtained. The alginic acid isolated from this brown seaweed was found to be predominantly of poly 1 → 4β-d-mannuronic acid type. The water-soluble sulphated polymer showed high anticoagulant activity.     

Transport of d-glucose by brush border membranes isolated from the renal cortex

The transport of d-glucose by brush border membranes isolated from the rabbit renal cortex was studied. At concentrations less than 2 mM, the rate of d-glucose uptake increased linearly with the concentration of the sugar. No evidence was found for a “high-affinity” (μM) saturable site. Saturation was indicated at concentrations of d-glucose greater than 5 mM. The uptake of d-glucose was stereospecific and selectively inhibited by d-galactose and other sugars. Phlorizin inhibited the uptake of d-glucose in the presence and absence of Na⁺. The glycoside was a potent inhibitor of the efflux of d-glucose. Preloading the brush border membrane vesicles with d-glucose, but not with l-glucose, accelerated exchange diffusion of d-glucose. These results demonstrate that the uptake of d-glucose by renal brush borders represents transport into an intravesicular space rather than solely binding. The rate of d-glucose uptake was increased when the Na⁺ in the extravesicular medium was high and the membranes were preloaded with a Na⁺-free medium. The rate of d-glucose uptake was inhibited by preloading the brush border membranes with Na⁺. These results are consistent with the Na⁺ gradient hypothesis for d-glucose transport in the kidney. Thus, the presence of a Na⁺-dependent facilitated transport of d-glucose in isolated renal brush border membranes is indicated. This finding is consistent with what is known of the transport of the sugar in more physiologically intact preparations and suggests that the membranes serve as an effective model system in examining the mechanism of d-glucose transport in the kidney.     

Composition and properties of the extracellular polysaccharide produced by arthrobacter stabilis NRRL B-3225

The extracellular polysaccharide produced by Arthrobacter stabilis NRRL B-3225 contains d-glucose, d-galactose, pyruvic acid, O-succinyl, and O-acetyl in the approximate molar ratio of 6:3:1:1:1.5. Succinyl is linked as its half-ester, making it a readily removable, acidic function that imparts versatility to the polysaccharide both for fundamental research and for commercial use. The viscosity of aqueous, salt-free solutions of both native and deacylated polymer is relatively low, but atypical of anionic polysaccharides, increases rapidly in the presence of salts, acids, or alkali.