Showing papers on "Paper chromatography published in 1974"

Covalent structure of calf-thymus ALK-histone.

Abstract: Highly purified calf thymus ALK-histone (histone rich in alanine, lecine and lysine) was isolated from F2a2 histone fraction by ion-exchange chromatography on Biorex 70 followed by gel filtration chromatography on Sephadex G-100 Peptides obtained by enzymatic hydrolyses (trypsin, chymotrypsin, thermolysin) of native or maleylated protein were fractionated by ion-exchange chromatography on Chromobeads P Thermolysin peptides which account for 127 of the 129 residues of the protein provide overlaps of tryptic and chymotryptic peptides and led us to establish the complete amino-acid sequence of ALK-histone as follows: Ser1Gly-Arg-Gly-Lys-Gln-Gly-Gly-Lys-Ala-10Arg-Ala-Lys-Ala- Lys-Thr-Arg-Ser-Ser-Arg20-Ala-Gly-Leu-Gln-Phe-Pro-Val-Gly-Arg-Val30-His-Arg-Leu-Leu-Arg-Lys-Gly-Asn-Tyr-Ala40-Glu-Arg-Val-Gly-Ala-Gly-Ala-Pro-Val-Tyr50-Leu-Ala-Ala-Val-Leu-Glu-Tyr-Leu-Thr-Al60-Glu-Ile-Leu-Glu-Leu-Ala-Gly-Asn-Ala-Ala70-Arg-Asp-Asn-Lys-Lys-Thr-Arg-Ile-Ile-Pro80-Arg-His-Leu-Gln-Leu-Ala-Ile-Arg-Asn-Asp90-Glu-Glu-Leu-Asn-Lys-Leu-Leu-Gly-Lys-Val100-Thr-Ile-Ala-Gln-Gly-Gly-Val-Leu-Pro-Asn110-Ile-Gln-Ala-Val-Leu-Leu-Pro-Lys-Lys-Thr120Glu-Ser-His-His-Lys-Ala-Lys-Gly-Lys129 The amino end is acetylated The sequence of ALK-histone is characterized by two hydro-phobic regions, the first from Val-43 to Ala-70 and the second from Val-100 to Pro-117 and presents many repetitive or analogous sequences The NH2-terminal sequence of ALK-histone is highly basic and shows a remarkable analogy with that of GRK-histone A cluster of five basic residues His-His-Lys-Ala-Lys-Gly-Lys is present at the COOH-terminal end

Isolation and Characterization of Oligosaccharides from Canine Submaxillary Mucin

Abstract: Pure mucin isolated from canine submaxillary glands was treated with alkaline borohydride and products of degradation were fractionated by a procedure that included ion-exchange chromatography, gel filtration, high-voltage electrophoresis and paper chromatography. Seven acidic, reduced oligosaccharides were isolated being divided into two distinct types: one, containing sialic acid and no glucosamine or sulfate (type A) and the other containing glucosamine and sulfate but no sialic acid (type B). The most complete oligosaccharide of type A designated component (I) was a tetrasaccharide α-l-fucopyranosyl-(1 2)-d-galactopyranosyl-[N-acetylneuraminyl-(2 6)]-2-acetamido-2-deoxy-d-galactitol. The most complex oligosaccharide of type B designated component (V) has a hexasaccharide with the following structure: α-l-fucopyranosyl-(1 2)-β-d-galactopyranosyl-(1 3, 4, or 6)-β-2-acetamido-2-deoxy-d-glucopyranosyl 3 or 4 sulfate-(1 6)-α-l-fucopyranosyl-(1 2)]-β-d-galactopyranosyl-(1 3) 2-acetamido-2-deoxy-d-galactitol. All the other acidic isolated oligosaccharides were derived from these two main components. No oligosaccharide containing both sialic acid and glucosamine sulfate could be demonstrated. Based on these structural features, a possible biosynthetic pathway leading to the two different oligosaccharide chains is discussed. Variations in ratios of sugars observed in canine submaxillary mucin and especially the reciprocal relationship between sialic acid and fucose noted earlier (1962) by Dische, Pallavicini, Kavasaki, Smirnow, Cizek and Chien may reflect variations in the proportions of these two types of oligosaccharides.

Lysis of Yeast Cell Walls: Glucanases from Bacillus circulans WL-12

Abstract: Endo-beta-(1 --> 3)- and endo-beta-(1 --> 6)-glucanases are produced in high concentration in the culture fluid of Bacillus circulans WL-12 when grown in a mineral medium with bakers' yeast cell walls as the sole carbon source. Much lower enzyme levels were found when laminarin, pustulan, or mannitol was the substrate. The two enzyme activities were well separated during Sephadex G-100 chromatography. The endo-beta-(1 --> 3)-glucanase was further purified by diethylaminoethyl-cellulose and hydroxyapatite chromatography, whereas the endo-beta-(1 --> 6)-glucanase could be purified further by diethylamino-ethyl-cellulose and carboxymethyl cellulose chromatography. The endo-beta-(1 --> 3)-glucanase was specific for the beta-(1 --> 3)-glucosidic bond, but it did not hydrolyze laminaribiose; laminaritriose was split very slowly. beta-(1 --> 4)-Bonds in oat glucan in which the glucosyl moiety is substituted in the 3-position were also cleaved. The kinetics of laminarin hydrolysis (optimum pH 5.0) were complex but appeared to follow Michaelis-Menten theory, especially at the lower substrate concentrations. Glucono-delta-lactone was a noncompetitive inhibitor and Hg(2+) inhibited strongly. The enzyme has no metal ion requirements or essential sulfhydryl groups. The purified beta-(1 --> 6)-glucanase has an optimum pH of 5.5, and its properties were studied in less detail. In contrast to the crude culture fluid, the two purified beta-glucanases have only a very limited hydrolytic action on cell wall of either bakers' yeast or of Schizosaccharomyces pombe. Although our previous work had assumed that the two glucanases studied here are responsible for cell wall lysis, it now appears that the culture fluid contains in addition a specific lytic enzyme which is eliminated during the extensive purification process.

The extracellular products of Anabaena cylindrica Lemm. I. Isolation of a macromolecular pigment-peptide complex and other components

Abstract: The extracellular products of Anabaena cylindrica Lemm. comprise a large variety of compounds including peptides, brownish pigments and substances fluorescing white and blue in ultraviolet light. A number were separated or isolated using techniques of gel filtration, ion exchange and paper chromatography. Serine and threonine comprised over 90% of the amino acids in a group of complex pigmented and fluorescent compounds. One of these accounted for a large proportion of the peptide and pigment present. It contained a large pigment moiety of molecular weight > 5 000 which formed a firm complex with more than 10% of the iron supplied in the culture medium. The anti-polymyxin activity described by Whitton was not associated with any of the major pigments or peptides present.

Role of the N-terminal amino acid for the biological activities of angiotensin and inhibitory analogues.

Abstract: Aspartic acid was replaced in position 1 of angiotensin II (ATII) with several amino acids, to assess the possible influence of the N-terminal amino acid for (a) the intrinsic activity, (b) the affinity, and (c) the metabolic degradation of agonist analogues of ATII. Some of the substitutions in position 1 were used in combination with replacement of Phe by Gly or Leu in position 8, to obtain the corresponding antagonist.The compounds were tested in vivo (rat blood pressure) and in two in vitro preparations (rat stomach and rabbit aorta strips). The oil immersion technique, described by Kalsner and Nickerson (1968) (Can. J. Physiol. Pharmacol. 46, 719–730), was used to study the disposition of the peptides by vascular smooth muscles (rabbit aorta strips). Degradation of the peptides by purified aminopeptidases was evaluated in vitro by measuring the fragments on paper chromatography. Potency of antagonists was estimated in vivo (ID50) and in vitro (pA2 values): duration of action was established by infusi...

Chemical evolution. XXI. The amino acids released on hydrolysis of HCN oligomers

Abstract: Major amino acids released by hydrolysis of acidic and basic HCN oligomers are identified by chromatography as Gly, Asp, and diaminosuccinic acid. Smaller amounts of Ala, Ile and alpha-aminoisobutyric acid are also detected. The amino acids released did not change appreciably when the hydrolysis medium was changed from neutral to acidic or basic. The presence of both meso and d, l-diaminosuccinic acids was established by paper chromatography and on an amino acid analyzer.

Lipoteichoic Acids from Streptococcus sanguis

Abstract: Two lipoteichoic acids, membrane (MLTA) and wall (WLTA), have been purified from Streptococcus sanguis by Sepharose and Ecteola-cellulose column chromatographies and concanavalin A-conjugated Sepharose affinity column chromatography. The teichoic acids were homogenous as judged by disc gel electrophoresis, column chromatography, and double diffusion tests. Both MLTA and WLTA consisted of glycerol, phosphate, glucose, and fatty acids in the ratios of 0.95:1:0.71:0.046 and 0.99:1:0.79:0.023, respectively. alpha-Glycerol-phosphate was obtained by the partial acid hydrolysis of the lipoteichoic acids suggesting that their backbone structure consists of the glycerol moieties linked by 1, 3-phosphodiester bonds. Both WLTA and MLTA form aggregates, perhaps due to micelle formation, in concentrated solution. The aggregate form of MLTA dissociates to a much greater extent than that of WLTA under similar conditions.

The Sequence of Sheep κ-Casein: Primary Structure of para-κA-Casein

Abstract: The primary structure of sheep para-xA-casein, the N-terminal portion of x-casein, was established. The reduced alkylated protein was subjected to digestion with trypsin. The resulting soluble peptides were purified by a combination of Dowex 1 X2 chromatography and electrophoresis and chromatography on paper; the core peptides were solubilized in 50% formic acid and filtered on Sephadex G-50. The amino-acid sequence of these peptides was determined chiefly with a Sequencer. Alignment of the tryptic peptides into a single chain containing 105 amino acids was determined from basic overlap peptides (tryptic core peptides with several basic amino acids; chymotryptic peptides). Some comparisons were achieved with cow para-xa-casein. %-Casein is the principal casein fraction affected by chymosin (or rennin) in the primary phase of the milk clotting process [ 11. The group of Jollbs established already in 1965 that during this enzymic reaction, a Phe -+ Met linkage was specifically split in cow as well as in sheep x-caseins [2-41. An insoluble part, para-x-casein (N-terminal moiety of x-casein), was from Merck or Prolabo except those employed for the Sequencer which were purchased from Socosi (94100Saint Maur, France) and 4-sulfophenylisothiocyanate from Pierce.

Methods for the Isolation and Determination of Glutamate, Glutamine, Aspartate, and γ-Aminobutyrate in Brain

Abstract: With the advent of various types of chromatography, a wide choice of methods is now available for the resolution and isolation of amino acids in complex mixtures. For quantitative analysis, paper chromatography and ion-exchange chromatography are the methods of choice. One-dimensional rather than two-dimensional paper chromatography is preferred as a routine procedure, samples being pretreated with ion-exchange resins to obtain groups of amino acids. Reliable methods for the isolation of the amino acids in the brain after paper chromatography have been published (Gaitonde, 1961; Gaitonde et al., 1965; Robinson and Williams, 1965).

Isolation and Characterization of Neutral Peptides in Soy Sauce

Abstract: A soy sauce sample was fractionated by gel filtration on a Sephadex G–15 column, then the fractions were subfractionated on the basis of acidity by ion exchange chromatography on a QAE-Sephadex A–25 column. After preliminary fractionation, the components in the neutral subfractions were transformed to copper salts, and the salts were chromatographed on a DEAE-Sephadex A–25 column with a borate buffer and aqueous acetic acid to separate neutral peptide substances from a large amount of free amino acids. The peptide fractions were further fractionated on a preparative amino acid analyzer and by paper chromatography to separate the peptide substances.Three glycodipeptides and eight dipeptides were isolated and characterized as the major neutral peptide components in soy sauce. However, it was difficult to anticipate any direct contribution of these peptides to the flavor construction in soy sauce on the basis of their taste intensities and contents.

The tryptic peptides of rabbit muscle triose phosphate isomerase.

Abstract: 1. The peptides obtained by tryptic digestion of S-[14C]carboxymethylated rabbit muscle triose phosphate isomerase have been studied. 2. The first step in the fractionation of the tryptic digest was gel filtration on coupled columns of Sephadex G-25 and G-50. Further fractionation was carried out by paper electrophoresis and paper chromatography. 3. The digest contained 26 peptides and three free amino acids. The sizes of the peptides ranged from two to 29 residues. 4. The sequences of the peptides have been determined. 5. The length of the polypeptide chains is about 250 amino acid residues. 6. The variant sequences encountered were due to partial deamidation; this may be one of the reasons for multiple forms of the enzyme. 7. The chicken and rabbit enzymes are compared. 8. Detailed evidence for the sequences of the tryptic peptides has been deposited as Supplementary Publication SUP 50024 at the British Library, Lending Division (formerly the National Lending Library for Science and Technology), Boston Spa, Yorks. LS23 7BQ, U.K., from whom copies can be obtained on the terms given in Biochem. J. (1973) 131, 5.

The fate of 2,6-dimethoxy(U-14C)phenol in the rat.

Abstract: 1. The metabolic fate of 2,6-dimethoxy[U-14C]phenol, administered intravenously to rats at three dose levels (10–30 mg/kg body wt.), was investigated.2. The majority of injected radioactivity appea...

Microbial production of vitamin b12 antimetabolites. i

Abstract: One of the vitamin B12 antimetabolites produced by Bacillus cereus 439 has been identified as N5-hydroxy-L-arginine on the basis of its mass-spectrum, 13C-NMR, and mobilities in thin-layer chromatography and paper chromatography in comparison with standard material. The inhibition of growth of Escherichia coli (Davis 113-3)by this compound was reversed by vitamin B12 and was potentiated by L-lysine and 4-hydroxy-L-lysine. Vitamin B12 also partially reversed the inhibition of growth by N5-hydroxy-L-arginine of Pseudomonas aeruginosa, other strains of E. coli, and Klebsiella pneumoniae. While neither N5-hydroxy-L-arginine nor L-lysine affected the growth in tissue culture of KB cells, the mixture of these two amino acids was quite inhibitory.

6‐O‐Methyl‐d‐glucosamine in Lipopolysaccharides of Rhodopseudomonas palustris Strains

Abstract: A hitherto unknown amino sugar was discovered in lipopolysaccharides isolated from strains of chemotype III of the photosynthetic bacterium Rhodopseudomonas palustris. The sugar (2–3% of the lipopolysaccharides dry weight) is detected as a shoulder on the descending part of the glucosamine peak obtained when acid hydrolysates of the lipopolysaccharides are passed through an amino acid analyzer. It was separated from the other amino sugars of the lipopolysaccharide (i.e. glucosamine, galactosamine, quinovosamine) by high-voltage paper electrophoresis and subsequent paper chromatography. The mass spectrometric analysis of the isolated sugar as deuterium-reduced alditol acetate revealed the presence of a 6-O-methyl-2-amino-2-deoxyhexose. Demethylation of the isolated sugar afforded glucosamine and the demethylation of its ninhydrin degradation product yielded arabinose, which showed that the parental amino sugar is glucosamine. Authentic 6-O-methyl-d-glucosamine exhibited identical properties in paper chromatography and high-voltage electrophoresis as did its alditol acetate derivative in gas-liquid chromatography and mass spectrometry. These data prove that the unknown amino sugar in lipopolysaccharides of Rhodopseudomonas palustris is 6-O-methyl-glucosamine. The strong positive optical rotation of the isolated sugar indicates its d-configuration. 6-O-Methyl-d-glucosamine is part of the O-specific chain and not of thelipid A moiety of the lipopolysaccharide. Periodate-oxidation studies revealed that the sugar, like the other amino sugars of this lipopolysaccharide, does not occupy non-reducing terminal positions to any appreciable extent.

Production of cephalosporin C by Paecilomyces persicinus P-10.

Abstract: After the growth of Paecilomyces persicinus P-10 in a glucose-peptone medium, filtrates were collected and analyzed for antibiotic antivity. Activities against Salmonella gallinarum ATCC 3030 and Alcaligenes faecalis ATCC 8750 (penicillin N-resistant strain) were obtained. Part of the former activity was readily inactivated by penicillinase. The fraction active against A. faecalis was isolated by passage through Amberlite XAD-2 and Amberlite IRA-68. The powder eventually obtained was subjected to paper chromatography followed by bioautography, and the activity obtained corresponded to that of a sample of cephalosporin C. Thin-layer chromatography was also employed to verify the presence of cephalosporin C in the P-10 powder. The active solids were further purified by means of paper chromatography in a solvent system consisting of n -butanol-acetic acid-water (60:15:25, vol/vol). The material obtained from this procedure yielded an infrared absorption spectrum identical to that of cephalosporin C. Similarly, the ultraviolet absorption of the purified preparation coincided with that of cephalosporin C. Exposure of the purified solids to cephalosporinase resulted in rapid inactivation of the antibiotic. In addition to penicillin N and cephalosporin C, filtrates of P. persicinus P-10 also contained deacetylcephalosporin C, deacetoxycephalosporin C, and cephalosporin P.

Properties of conjugation hormones (erogens) from the basidiomycete Tremella mesenteria

Abstract: Hormones produced by haploid cells of one mating type of T. mesenterica and inducing conjugation tubes in the other mating type have been studied in preparation for isolation and identification. The active material can be extracted from aqueous solution with n-butanol, but not with less polar solvents, and is adsorbed on cation and anion exchange resins, charcoal, and neutral polystyrene resin. The molecular weight is probably less than 1000. Three active components can be separated by silica gel column chromatography with a gradient of water in ethanol, or by paper chromatography. The conjugation hormones may be non-polar amino acids or small peptides.

Metabolism of Trehalose in Euglena gracilis

Abstract: The role of β-glucose 1,6-bisphosphate as a cofactor in the reaction catalyzed by the phosphoglucomutase for β-glucose 1-phosphate (β-phosphoglucomutase) has been examined in purified extracts of Euglena gracilis var. bacillaris. 1 The incubation of β-glucose 1-[32P]phosphate with β-phosphoglucomutase in presence of high concentrations (0.1 mM) of a commercial preparation of α-d(+)-glucose 1,6-bisphosphate (prepared synthetically) yielded a labelled compound running electrophoretically and chromatographically as the sugar bisphosphate. Specificity studies with β-phosphoglucomutase and muscle phosphoglucomutase (α-phosphoglucomutase) strongly suggest that the compound formed is β-glucose 1,6-[32P]bisphosphate. The results would also indicate that the commerical preparation of α-d(+)-glucose 1,6-bisphosphate contains β-glucose 1,6-bisphosphate as contaminant. 2 When synthetic β-glucose 1-phosphate preparations were purified by paper chromatography and then incubated with β-phosphoglucomutase, no glucose 6-phosphate could be detected. On the other hand the reaction readily took place when challenged with the chromatographic fraction that runs as sugar bisphosphate. Upon analysis with electrophoresis, chromatography and weak acid hydrolysis, and experiments with β-phosphoglucomutase and α-phosphoglucomutase it was concluded that this compound is β-glucose 1,6-bisphosphate. 3 It was established that the natural α-d(+)-glucose 1,6-bisphosphate not only fails to sustain the reaction catalyzed by β-phosphoglucomutase but rather inhibits the reaction when added to the whole β-phosphoglucomutase system. It was also demonstrated that β-glucose 1,6-bisphosphate acts as inhibitor of the α-phosphoglucomutase system. 4 These results show for the first time that β-glucose 1,6-bisphosphate is an active and necessary participant in the reaction catalyzed by β-phosphoglucomutase. The role played by β-glucose 1,6-bisphosphate in this reaction would be essentially the same as that of the α-anomer in the reaction catalyzed by α-phosphoglucomutase.

Identification d'une cytokinine par chromatographic en phase gazeuse à partir de cultures pures de Taphrina cerasi

Abstract: The ascomycete Taphrina cerasi (Fuck.) Sad., which induces malformations known as "witches’ brooms," when cultured in liquid media releases cytokinins. One of these, isolated from culture filtrates by ethyl acetate extraction, was identified as zeatin by paper chromatography, a biological test, and gas-liquid chromatography.