Analysis of Monosaccharides

Manzi, Adriana E.; Powell, Leland D.; Varki, Ajit

doi:10.1002/0471142727.mb1718s32

Cited by 4 publications

(1 citation statement)

References 32 publications

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“…A sample of polysaccharide (5 mg) was hydrolyzed by HCl (2 N) at 100°C (4 h). The monosaccharide composition of the acid-hydrolysis products was determined by HPLC on a carbohydrate analyzer (Biotronik, ISA-07/S2504 column, 0.4 u 25 cm, Shimadzu) with detection by the bicinchoninate method [24].…”

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Structural characteristics and antitumor activity of a new chrysolaminaran from the diatom alga Synedra acus

et al. 2010

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The 1o3;1o6-E-D-glucan chrysolaminaran with molecular weight 8.5 kDa and 3.6:1 ratio of 1o3:1o6 bonds was isolated from cultivated diatom alga Synedra acus and characterized. It was shown that the isolated chrysolaminaran inhibited growth and colony formation of human colon tumor cells.Keywords: diatom alga, chrysolaminaran, 1o3;1o6-E-D-glucan, antitumor activity. Diatom algae are unicellular or colonial eukaryotic organisms that inhabit seas, oceans, and freshwaters and have adapted to habitats of plankton and benthos. Diatom algae and terrestrial plants are responsible for about 40% of the primary production on Earth [1]. Polysaccharides make up from 10 to 80% of the organic mass of diatoms [2, 3] and play an important role in their metabolism. Polysaccharides of diatom algae are divided into three groups depending on their location in the cell: storage, structural, and extracellular [4,5]. Each of these fulfills its own specific function.Storage polysaccharides are localized in vacuoles and represent the final photosynthesis products [6]. They are E-D-glucans, are called chrysolaminarans, and consist of short chains of 1o3-bonded E-D-glucopyranoses (n = 20-60) with branching at C6 and/or C2 [7-9].The study of polysaccharides from diatom algae is important for both an understanding of the functional processes of these organisms and their practical use. Analogs of chrysolaminarans, 1o3;1o6-E-D-glucans, isolated from yeast, fungi (lentinan), and culture filtrates of Schizophyllum commune (schizophyllan) have been used in cancer immunotherapy [10,11], act as immunomodulators, and increase the resistance to infections [12]. This makes them an attractive alternative to cytotoxic drugs. In contrast with other natural biopolymers, purified E-1,3-glucans retain their biological activity over time so that the mechanism of their action on the cellular and molecular level can be studied. It is currently known that the biological action of E-1,3-glucans is mediated through two membrane receptors, CR3 [13] and Dectin-1 [14].The biological activity of algae polysaccharides (laminarans and chrysolaminarans) has been studied less than that of glucans from yeast and fungi.Our goal was to isolate chrysolaminaran from cultivated diatom alga Synedra acus and to study its structure and antitumor activity.A chrysolaminaran preparation consisting of a polysaccharide-protein complex with a 1:8 protein:carbohydrate ratio was isolated from S. acus cultivated under laboratory conditions by extraction with hot water. According to the literature, the protein:carbohydrate ratio in similar complexes can vary from 0.33 to 6.5 depending on the physiological condition of the cell [15]. Removal of protein from the S. acus complex by the Sevage method [16] did not completely free the polysaccharide from the protein. A sample of the chrysolaminaran without protein could be obtained using hydrophobic chromatography over Polykhrom-1. The chrysolaminaran yield was 0.35% of the moist alga weight.

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Structural characteristics and antitumor activity of a new chrysolaminaran from the diatom alga Synedra acus

et al. 2010

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Purification and characterization of an adhesin from Pasteurella haemolytica

Jaramillo

Dı́az²,

Hernández³

et al. 2000

Glycobiology

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We purified an adhesin from Pasteurella. haemolytica by affinity chromatography using glutaraldehyde treated rabbit erythrocytes stroma. The adhesin is a protein of 68 kDa, as determined by SDS-PAGE, and the most abundant amino acids constituting this protein were Gly, Ser, Glx, and Ala, and low concentrations of Cys, Met, and Tyr residues were also found. The N-terminal sequence of the adhesin is ANEVNVYIYKQPYLI. No carbohydrate residues were detected. The adhesin agglutinated rabbit erythrocytes but when the latter were desialylated or pronase treated the agglutinating activity was abolished. The agglutinating activity of the adhesin was inhibited with N-acetyl-D-glucosamine (GlcNAc), and in a lesser degree with N-acetyl-neuraminic acid (NeuAc). GalNAc, N-glycolyl-neuraminic acid, N-deacetylated GlcNAc, or neutral sugars do not modify the activity of the adhesin. The equatorial -OH on C4 and the NH-acetylated group on C2 from GlcNAc, as well as the 4-OH and NH-acetylated group on C5 from NeuAc seem to be responsible for the interaction with the adhesin. The protein is divalent cation-dependent and thermolabile. As for the agglutinating activity, the adhesion of P.haemolytica to tracheal cell-cultures was inhibited by GlcNAc, NeuAc or the purified adhesin, strongly suggesting that the P.haemolytica adhesin plays an important role in infection.

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Use of inhibitors to characterize intermediates in the processing of N- glycans synthesized by insect cells: a metabolic study with Sf9 cell line

et al. 1999

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The most frequent type of N-glycan synthesized by lepidopteran Sf9 cells appears to be fucosylated Man3GlcNAc2,and this has been a limitation for a large scale production and utilization of therapeutic glycoproteins in cultured insect cells. The current knowledge of the protein glycosylation pathway derived from structural studies on recombinant glyco-proteins expressed by using baculovirus vectors. In this work we provide more direct evidence for the sequential events occurring in the processing of endogenous N-glycoproteins of noninfected Sf9 cells. By metabolic labeling with radioactive mannose, we characterized the glycan structures which accumulated in the presence of processing inhibitors (castanospermine and swainsonine) and in the presence of an intracellular trafficking inhibitor (monensin). We thus demonstrated that from the glycan precursor Glc3Man9GlcNAc2 to GlcNAcMan5(Fuc)GlcNAc2 intermediate, the processing pathway in Sf9 cells paralleled the one demonstrated in mammalian cells. By using monensin, we demonstrated the formation of Man3(Fuc)GlcNAc2 from GlcNAcMan3(Fuc)GlcNAc2, a reaction which has not been described in mammalian cells. Our results support the idea that the hexosaminidase activity is of physiological relevance to the glycosylation pathway and is Golgi located.

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Analysis of Monosaccharides

Cited by 4 publications

References 32 publications

Structural characteristics and antitumor activity of a new chrysolaminaran from the diatom alga Synedra acus

Structural characteristics and antitumor activity of a new chrysolaminaran from the diatom alga Synedra acus

Purification and characterization of an adhesin from Pasteurella haemolytica

Use of inhibitors to characterize intermediates in the processing of N- glycans synthesized by insect cells: a metabolic study with Sf9 cell line

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