Showing papers on "Melibiose published in 1994"

Carbon Source Requirements for Exopolysaccharide Production by Lactobacillus casei CG11 and Partial Structure Analysis of the Polymer.

Abstract: Exopolysaccharide production by Lactobacillus casei CG11 was studied in basal minimum medium containing various carbon sources (galactose, glucose, lactose, sucrose, maltose, melibiose) at concentrations of 2, 5, 10, and 20 g/liter. L. casei CG11 produced exopolysaccharides in basal minimum medium containing each of the sugars tested; lactose and galactose were the poorest carbon sources, and glucose was by far the most efficient carbon source. Sugar concentrations had a marked effect on polymer yield. Plasmid-cured Muc- derivatives grew better in the presence of glucose and attained slightly higher populations than the wild-type strain. The values obtained with lactose were considerably lower for both growth and exopolysaccharide yield. The level of specific polymer production per cell obtained with glucose was distinctively lower for Muc- derivatives than for the Muc+ strain. The polymer produced by L. casei CG11 in the presence of glucose was different from that formed in the presence of lactose. The polysaccharide produced by L. casei CG11 in basal minimum medium containing 20 g of glucose per liter had an intrinsic viscosity of 1.13 dl/g. It was rich in glucose (76%), which was present mostly as 2- or 3-linked residues along with some 2,3 doubly substituted glucose units, and in rhamnose (21%), which was present as 2-linked or terminal rhamnose; traces of mannose and galactose were also present.

Characterization of Eubacterium coprostanoligenes sp. nov., a Cholesterol-Reducing Anaerobe?

Abstract: A small, anaerobic, gram-positive coccobacillus that reduces cholesterol to coprostanol was isolated from a hog sewage lagoon. This isolate, strain HLT (T = type strain) does not require cholesterol for growth, but it requires lecithin and has phospholipase activity. Much acid is produced by the fermentation of amygdalin, lactose, and salicin. Arabinose, cellobiose, fructose, glucose, mannose, and melibiose are fermented weakly. Acetic, formic, and succinic acids are produced, as is hydrogen. The isolate does not reduce nitrate, produce indole, or hydrolyze starch and gelatin. Esculin is hydrolyzed. The properties of strain HLT are most similar to those of members of the genus Eubacterium. Because strain HL (= ATCC 51222) has unique morphological and physiological properties, we propose that it should be the type strain of a new species in the genus Eubacterium, Eubacterium coprostanoligenes.

Hydrolysis of galactooligosaccharides by commercial preparations of α‐galactosidase and β‐fruetofuranosidase: Potential for use as dietary additives

Abstract: In vitro and in vivo studies were conducted with six commercial enzyme preparations (SP249, Energex, Rohament CW, Novozyme 230 and crude α -galactosidase) to determine their effectiveness in hydrolysing galactooligosaccharides from soya bean and canola meal in the gastrointestinal tract of poultry. The use of the enzyme invertase to enhance galactoside hydrolysis was also studied. A wide range of α -galactosidase activity was observed in vitro, with crude α-galactosidase from Mortirella vinacea and Novozyme 230 preparation showing the highest activity values of 4.3 and 1.5 nkat mg−1, respectively. All preparations with the exception of crude α-galactosidase showed invertase activity which is known to convert raffinose and stachyose to the corresponding di-and trisaccharide, melibiose and manninotriose. Although the activity of invertase was highest on sucrose, the Novozyme 230 preparation showed activity values of 4.2 and 2.3 nkat mg−1 toward raffinose and stachyose substrates, respectively. De novo synthesis of raffinose was observed when soya bean meal, canola meal or pure sucrose and galactose were incubated with certain enzyme preparations (ie Energex). In general, preparations possessing hydrolytic activity towards galactooligosaccharides showed very little synthesis of raffinose while preparations capable of generating raffinose were very weak in the hydrolysis of galactooligosaccharides. The best result in terms of galactooligosaccharide in vitro hydrolysis of canola and soya bean meal was obtained with a combination of α-galactosidase and invertase. In the in vivo study with caecectomised hens, hydrolysis of galactooligosaccharides averaged 88% when crude α-galactosidase (2 g kg−1) and invertase (1 g kg−1) were added to laying, hen diet containing 200 g soya bean meal per kilogram. A problem identified in the current study was that minerals such as calcium phosphate and calcium carbonate common in poultry diets inhibit the hydrolysis activity of α-galactosidase, indicating that high levels of activity would be required to yield a response in practical poultry feeding.

Regulation of the raffinose permease of Escherichia coli by the glucose-specific enzyme IIA of the phosphoenolpyruvate:sugar phosphotransferase system.

Abstract: In enteric bacteria, chromosomally encoded permeases specific for lactose, maltose, and melibiose are allosterically regulated by the glucose-specific enzyme IIA of the phosphotransferase system. We here demonstrate that the plasmid-encoded raffinose permease of enteric bacteria is similarly subject to this type of inhibition.

Positive selection for resistance to 2-deoxyglucose gives rise, in Streptococcus salivarius, to seven classes of pleiotropic mutants, including ptsH and ptsI missense mutants.

Abstract: Summary We have used the toxic non-metabolizabie glucose/ mannose analogue 2-deoxygiucose to isolate a comprehensive collection of mutants of the phosphoenoipyruvate:sugar phosphotransferase system from Streptococcus salivarius. To increase the range of possible mutations, we isolated spontaneous mutants on different media containing 2-deoxyglucose and various metabolizable sugars, either lactose, meli-biose, galactose or fructose. We found that the frequency at which 2-deoxygiucose-resistant mutants Were isolated varied according to the growth substrate. The highest frequency was obtained with the combination galactose and 2-deoxygiucose and was 15-fold higher than the rate observed with the mixture melibiose and 2-deoxygiucose, the combination that gave the lowest frequency. By combining results from: (i) Western biol analysis of IIIMan, a specific component of the phosphoenolpyruvate:mannose phosphotransferase system in S. salivarius; (ii) rocket immunoelectrophoresis of HPr and EI, the two general energy-coupling proteins of the phosphotransferase system; and (iii) from gene sequencing, mutants could be assigned to seven classes. Class 1 was composed of strains devoid of IIIManL, a low-molecular-weight form of IIIManL (35200), class 2 was composed of strains exhibiting a reduced level of IIIManL, class 3 was composed of strains devoid of both forms of IIIMan (IIIManL as well as IIIManH, the high-molecular-weight form of IIIMan (38900)), class 4 was composed of mutants bearing a mutation in ptsH, the gene encoding HPr, class 5 was composed of mutants bearing a mutation in ptsl, the gene encoding EI, class 6 was composed of 2-deoxygiucose-resistant strains without any apparent defect in PTS components, and class 7 was composed of strains possessing both forms of IIIMan but abnormal levels of HPr and/or EI without any mutation in the ptsH and/or the ptsI genes. Preliminary characterization of representative strains of each class is reported.

Human natural antibodies to porcine platelets.

Abstract: The specificity of human natural antibodies directed against blood cells from pigs was investigated by ELISA and immunoblotting. Both IgG and IgM were identified as xenoantibodies reacting with pig platelets adsorbed to microplates. The antibodies could be absorbed on platelets as well as on RBC, suggesting that the corresponding antigens are expressed on the surface of a variety of cells. Galactose (20 mM) and melibiose (10 mM) partially inhibited (approximately 50%) the binding of antibodies to platelets, whereas lactose and cellobiose (300 mM) did not. On immunoblots, platelet glycoproteins of 115, 125, 135, 180, and 210 kDa were specifically revealed with human sera diluted 1/20. In contrast with the results obtained by ELISA, xenoantibodies reactive with blotted glycoproteins were of the IgM class and the binding was not significantly inhibited by galactose or melibiose. "Anti-Gal" antibodies, purified from human sera by affinity chromatography on a melibiose-Sepharose immunoabsorbent, represented only a minor portion of the antibodies reactive with porcine platelets. Purified anti-Gal antibodies bound to the 115- and 135-kDa components, whereas the antibodies in the nonretained fraction revealed the 125-kDa molecule. As deduced from these data, human serum contains natural antibodies of both IgG and IgM classes directed to several porcine antigens. Gal-reactive structures were identified on the 115- and 135-kDa platelet glycoproteins, which might be homologous to their counterpart on endothelial cells. Also, the present work suggests that a majority of the natural antibodies reacted with other unidentified structures.

Escherichia coli heat-labile enterotoxin binds to glycosylated proteins with lactose by amino carbonyl reaction.

Abstract: The binding of Escherichia coli heat-labile enterotoxin (LT) type I to glycosylated proteins with lactose (Galβ1-4Glc) by amino carbonyl reaction was studied by the Western blot assay and by the microtiter well binding assay. LT bound to a lactose-α-lactalbumin amino carbonyl product (Lac-LA), whereas cholera toxin did not. The binding ability of Lac-LA was abolished by β-galactosidase treatment, indicating that the terminal galactose is essential for the binding of LT. The binding of LT to Lac-LA was inhibited by galactose and lactose, and most effectively inhibited by lactulose (Galβ1-4Fru), which is a structural analog of the Amadori rearrangement product of the amino carbonyl reaction between lactose and an e-amino group of a lysine residue (lactuloselysine). The results suggest that LT recognizes the portion of lactuloselysine in Lac-LA. LT also bound to a melibiose (Galα1-6Glc)-α-lactalbumin amino carbonyl product (Mel-LA), but the binding ability of Mel-LA was weaker than that of Lac-LA, suggesting that the β1-4 linked terminal galactose is dispensable but preferable for the binding. Furthermore, LT bound to the amino carbonyl products of lactose with β-lactoglobulin, caseins, bovine serum albumin, and ovalbumin. These results indicate that LT binds to the amino carbonyl products between proteins and sugars containing the terminal galactose, such as lactose.

Ultrasound-induced scission of the glycosidic bond in disaccharides

Abstract: The products originating from the scission of the glycosidic bond by ultrasound were investigated for lactose (4-O-β-d-galactopyranosyl-d-glucopyranose), melibiose (6-O-α-d-galactopyranosyl-d-glucopyranose) and trehalose (1,1-O-α,α-d-glucopyranosyl-d-glucopyranose). A comparison with the product formation of radiolysis reveals that for ultrasound the reactions also start from sugar radicals, produced by abstraction of carbon-bonded hydrogens by OH- and H-radicals. From these radicals, only those that are directly located at this bond or can be transferred by rearrangement into a position adjacent to this bond contribute to the scission of the glycosidic bond: C-1′, C-5′, C-4 for lactose; C-1′, C-5′, C-6 for melibiose and C-1 (=C-1′), C-5 (=C-5′) for trehalose. Based on the yields of the characteristic products for the different radicals it can be concluded that the reaction predominantly starts from the radical at C-1′ for lactose and melibiose and from C-1 (=C-1′) for trehalose. From the product formation it can be further deduced that the radical-induced scission is mainly determined by hydrolytic processes, fragmentation by radical rearrangement being of minor importance. The ratio of hydrolysis to fragmentation reactions reveals that in sonolysis this is still more pronounced than in radiolysis (lactose: sonolysis 8.2, radiolysis 4.2; melibiose: sonolysis 23.5, radiolysis 16.2).

Maltose transport in Aeromonas hydrophila: purification, biochemical characterization and partial protein sequence analysis of a periplasmic maltose-binding protein

Abstract: A clinical isolate of Aeromonas hydrophila was demonstrated to transport [14C]maltose with similar kinetics to enteric bacteria (Km: 0·3 μM; Vmax: 22 nmol min-1 per 109 cells). The uptake of [14C]maltose was completely inhibited in the presence of unlabelled maltose or maltodextrins, whereas other mono- and disaccharides, such as glucose, galactose, sucrose, lactose or melibiose, had no effect. A protein with an apparent molecular mass of 39 kDa (maltose-binding protein; MBP) was identified in osmotic-shock fluid of maltose-grown cells by SDS-gel electrophoresis, and was purified to homogeneity by either amylose affinity chromatography or ion-exchange chromatography. Equilibrium dialysis experiments revealed the ability of the purified protein to bind [14C]maltose with high affinity (KD = 1·6 μM). Unlabelled maltose and maltodextrins competed for the binding site. In a reconstitution experiment, A. hydrophila MBP poorly restored the transport activity of a binding-protein-deficient Escherichia coli (ΔmalE) mutant. N-terminal sequence analyses of the purified native protein and of peptides generated by cleavage with CNBr and subsequently separated by HPLC revealed about 56% identical amino acid residues, as compared to enterobacterial MBPs. We conclude that maltose is transported into A. hydrophila via a binding-protein-dependent transport system.

Transcriptional attenuation and differential mRNA stability in the regulation of the Escherichia coli melibiose operon.

Abstract: The organization of the melibiose operon of Escherichia coli is promoter-melA-melB. The amount of the product (alpha-galactosidase) of the first gene (melA) is much larger than that of the product (melibiose permease) of the second gene (melB). Using the chloramphenicol acetyltransferase gene (cat) as reporter, we found that there was an element between melA and melB, which reduced the expression of the downstream gene, melB. This region contained a boxA-like sequence, which is known as a binding site for an attenuation factor, NusA. Northern hybridization analysis revealed that the ratio of melA mRNA and melAB mRNA was comparable with the ratio of the melA and melB products. We also found that the melA mRNA was about 3-fold more stable than the melAB mRNA. Experimental results obtained with a nusAts mutant suggested that the NusA protein is involved in the reduced expression of the melB gene. We conclude that the production ratio of alpha-galactosidase and melibiose permease is regulated at two levels: 1) transcription and 2) mRNA stability.

Carbon requirements of Colletotrichum gloeosporioides Penz

Abstract: Monosporic isolates of Colletotrichum gloeosporioides were made from Punica grantum and one suitable isolate was selected for further work on carbon nutrition. Preliminary experiments showed that a pH value of 5, a temperature of 32°C and a period of 14 days were optimum conditions for the growth of this pathogen. Out of 41 carbon compounds tested, the pathogen showed excellent growth on starch, maltose, melibiose, dextrose, sucrose, raffinose, and dulcitol; good on tartaric acid, mannose, galactose, fructose, mannitol, and castor oil; fair on inulin, isopropyl alcohol, coconut oil, and pectin; poor on sorbose, n-butyl alcohol, arabionose, maleic acid, ethyl alcohol, succinic acid, citric acid, ribose, and malic acid, and no growth on the rest of the carbon compounds. In general, compounds which supported the best mycelial growth, yielded excellent or good sporulation of C. gloeosporioides and vice versa.

Building Blocks for Glycopeptide Synthesis. Disaccharide Glycosyl Isothiocyanates.

Abstract: The reaction of aldose oligosaccharides with aqueous ammonium hydrogencarbonate was revised and optimized for gram scale preparation of glycosylamines, using lactose, cellobiose, maltose, and melibiose as model compounds. The unprotected glycosylamines 1b-4b were transformed into the corresponding hepta-O-acetyl hydrochlorides 1e-4e via diethyl N-glycosylaminomethylenemalonates (1c-4c and 1d-4d). Reaction of le-3e with thiophosgene in a three-phased system led to the target hepta-O-acetylglycosyl isothiocyanates lf-3f in excellent yields. Under the same conditions, the melibiosyl derivative 4e underwent partial hydrolysis to give a mixture of melibiosyl isothiocyanate 4f and the reducing sugar derivative 4h. Treatment of 1f-4f with ammonia resulted in the quantitative formation of N-glycosylthioureas (1g-4g).