scholarly journals Isolation and characterization of sulphated mucopolysaccharides from rat leukaemic (RBL-1) basophils

1980 ◽  
Vol 185 (2) ◽  
pp. 367-372 ◽  
Author(s):  
D D Metcalfe ◽  
S I Wasserman ◽  
K F Austen
Keyword(s):  
Mast Cell ◽  
Salt Concentration ◽  
Tumour Cells ◽  
High Salt ◽  
High Salt Concentration ◽  
Dermatan Sulphate ◽  
Chondroitinase Abc ◽  
Isolation And Characterization

Proteoglycans of 300 000 mol.wt. were isolated from dispersed rat basophil tumour cells after labelling of the sulphated mucopolysaccharides with 35S in vitro:90% of the 35S-labelled mucopolysaccharides were extracted at high salt concentration. Alkali degradation of the 35S-labelled proteoglycans yielded glycosaminoglycan chains of 40 000 mol.wt. The composition of the salt-extracted 35S-labelled mucopolysaccharides, as defined by parallel or sequential degradation with chondroitinase AC, chondroitinase ABC and heparinase and resolution of the disaccharide-digestion products obtained with chondroitinase AC, was 48-61% chondroitin 4-sulphate, 20-30% dermatan sulphate, 10-15% heparin and 7-9% chondroitin 6-sulphate. Most of the salt-extracted 35S-labelled mucopolysaccharides were highly charged, with heparin and chondroitin 6-sulphate being relatively uniform in this regard, whereas chondroitin 4-sulphate and dematan sulphate exhibited a range of charge characteristics. The diversity of sulphated mucopolysaccharides present in the rat leukaemic basophil is in contrast with the predominance of heparin in the rat mast cell.

scholarly journals The copolymeric structure of pig skin dermatan sulphate. Isolation and characterization of l-idurono-sulphate-containing oligosaccharides from copolymeric chains

1974 ◽  
Vol 143 (2) ◽  
pp. 379-389 ◽  
Author(s):  
Lars-Åke Fransson ◽  
Lars Cöster ◽  
Birgitta Havsmark ◽  
Anders Malmström ◽  
Ingrid Sjöberg
Keyword(s):  
Periodate Oxidation ◽  
Exchange Chromatography ◽  
Dermatan Sulphate ◽  
Chondroitinase Abc ◽  
Pig Skin ◽  
Isolation And Characterization ◽  
Smith Degradation ◽  
Iduronic Acid ◽  
Acidic Conditions

Dermatan sulphate was degraded by testicular hyaluronidase and an oversulphated fraction was isolated by ion-exchange chromatography. This preparation, which contained fairly long segments derived from the non-reducing terminal portion of the molecule, was subjected to periodate oxidation under acidic conditions. The oxidized iduronic acid residues were cleaved by reduction-hydrolysis (Smith-degradation) (Fransson & Carlstedt, 1974) or by alkaline elimination. The oligosaccharides so obtained contained both GlcUA (glucuronic acid) and IdUA-SO4 (sulphated iduronic acid) residues. Copolymeric oligosaccharides obtained after alkaline elimination were cleaved by chondroitinase-AC into disaccharide and higher oligosaccharides. Since the corresponding oligosaccharides obtained by Smith-degradation were unaffected by this enzyme, it was concluded that the carbohydrate sequences were GalNAc-(IdUA-GalNAc)n-GlcUA-GalNAc. The iduronic acid-containing sequences were resistant to digestion with chondroitinase-ABC. It was demonstrated that the presence of unsulphated N-acetylgalactosamine residues in these sequences could be responsible for the observed effect. This information was obtained in an indirect way. Chemically desulphated dermatan sulphate was found to be a poor substrate for the chondroitinase-ABC enzyme. Moreover, digestion with chondroitinase-ABC of chondroitinase-AC-degraded dermatan sulphate released periodate-resistant iduronic acid-containing oligosaccharides. It is concluded that copolymeric sequences of the following structure are present in pig skin dermatan sulphate: [Formula: see text] N-acetylgalactosamine moieties surrounding IdUA-SO4 residues are unsulphated to a large extent.

scholarly journals Characterization of the RNA Synthesizing Activity of Isolated Kidney Nuclei

1975 ◽  
Vol 28 (6) ◽  
pp. 465
Author(s):  
CJ Story ◽  
J FW heldrake
Keyword(s):  
Activation Energy ◽  
Salt Concentration ◽  
Rna Synthesis ◽  
High Salt ◽  
High Salt Concentration ◽  
Limiting Factor ◽  
Isolated Kidney ◽  
Exogenous Dna ◽  
Low Salt

The limiting factor in RNA synthesis by isolated kidney nuclei is RNA nucleotidyltransferase at high salt concentrations but at low salt concentrations template availability becomes limiting. oc-Amanitin inhibits 85 % of the activity at high salt concentrations but only 20-50 % of the activity at low salt concentrations. Exogenous DNA is utilized at low salt concentrations [up to O�lM (NH4hS041 but not at high salt concentrations. The effect of increasing salt concentration is mainly to cause an increase in the length of chains synthesized. Initiation rates are not increased by high salt concentrations. The apparent Km for UTP is 8-10 11M at high salt concentrations, indicating that assays performed at low UTP concentrations are likely to give inaccurate results. The activation energy for the reaction at low salt concentration is less than that for the reaction at high salt concentration. The RNA synthesizing capacity of kidney nuclei is dependent on the method of isolation, and preparation by a modification of the Chauveau method (Chauveau et al. 1956) yields the most active nuclei.

PENGEMBANGAN TEKNOLOGI SELEKSI IN VITRO BERULANG (REPEAT CYCLING–IN VITRO SELECTION) TOLERAN STRES KEKERINGAN UNTUK PERBAIKAN MUTU GENETIK DAN PRODUKSI TANAMAN KEDELAI BERMIKORIZA

2017 ◽  
Vol 1 (1) ◽  
pp. 74-84
Author(s):  
Ahmad Riduan ◽  
Rainiyati Rainiyati ◽  
Yulia Alia
Keyword(s):  
Arbuscular Mycorrhizae ◽  
In Vitro Selection ◽  
Soil Samples ◽  
Single Spore ◽  
Isolation And Characterization ◽  
Single Spore Culture ◽  
Spore Culture ◽  
Glomus Sp

Every plant rhizospheres in any ecosystem there are various living microorganisms including Arbuscular Mycorrhizae Fungi (AMF).  An isolation and characterization is required to investigate the species or type of the AMF. This research was aimed at studying the isolation and characterization of AMF sporulation in soybean rhizospheres in Jambi Province. The results of evaluation on soil samples before trapping showed that there are spores from three genus of AMF twelve types Glomus , two types Acaulospora and one type of Enthrophospora.  Following single spore culture in soybean rhizosphere, 5 spore types were obtained:  Glomus sp-1, Glomus sp-4, Glomus sp-7, Glomus sp-8 Glomus sp-10.

Sand priming improves alfalfa germination under high-salt concentration stress

2006 ◽  
Vol 34 (1) ◽  
pp. 199-204 ◽  
Author(s):  
J. Hu ◽  
X.J. Xie ◽  
Z.F. Wang ◽  
W.J. Song
Keyword(s):  
Salt Concentration ◽  
High Salt ◽  
High Salt Concentration

scholarly journals Chondroitinase ABC digestion of dermatan sulphate. N.m.r. spectroscopic characterization of the oligo- and poly-saccharides

1989 ◽  
Vol 257 (2) ◽  
pp. 347-354 ◽  
Author(s):  
P N Sanderson ◽  
T N Huckerby ◽  
I A Nieduszynski
Keyword(s):  
Gel Permeation Chromatography ◽  
Spectroscopic Characterization ◽  
Repeat Sequence ◽  
Chemical Shift Assignments ◽  
Dermatan Sulphate ◽  
Chondroitinase Abc ◽  
Experimental Conditions ◽  
Full Analysis ◽  
Unsaturated Uronate

Dermatan sulphates, in which iduronate was the predominant uronate constituent, were partially digested by chondroitinase ABC to produce oligosaccharides of the following structure: delta UA-[GalNAc(4SO3)-IdoA]mGalNAc(4SO3) [where m = 0-5, delta UA represents beta-D-gluco-4-enepyranosyluronate, IdoA represents alpha-L-iduronate and GalNAc(4SO3) represents 2-acetamido-2-deoxy-beta-D-galactose 4-O-sulphate], which were fractionated by gel-permeation chromatography and examined by 100 MHz 13C-n.m.r. and 400/500 MHz 1H-n.m.r. spectroscopy. Experimental conditions were established for the removal of non-reducing terminal unsaturated uronate residues by treatment with HgCL2, and reducing terminal N-acetylgalactosamine residues of the oligosaccharides were reduced with alkaline borohydride. These modifications were shown by 13C-n.m.r. spectroscopy to have proceeded to completion. Assignments of both 13C-n.m.r. and 1H-n.m.r. resonances are reported for the GalNAc(4SO3)-IdoA repeat sequence in the oligosaccharides as well as for the terminal residues resulting from enzyme digestion and subsequent modifications. A full analysis of a trisaccharide derived from dermatan sulphate led to the amendment of published 13C-n.m.r. chemical-shift assignments for the polymer.

Arginine synthesis by hepatomas in vitro. II. Isolation and characterization of Morris hepatoma variants unable to convert ornithine to arginine, and modulation of urea-cycle enzymes by dexamethasone and cyclic-AMP

1984 ◽  
Vol 68 (1) ◽  
pp. 305-319
Author(s):  
S.J. Goss
Keyword(s):  
Cyclic Amp ◽  
Liver Cells ◽  
Isolation And Characterization ◽  
Rat Liver Cells ◽  
Morris Hepatoma ◽  
Ornithine Transcarbamoylase ◽  
Cellular Incorporation

‘77orn’, a derivative of the Morris rat hepatoma 7777, stably expresses high levels of ornithine transcarbamoylase (OTC) and carbamoylphosphate synthetase I (CPS-I), and is able to grow indefinitely in ornithine-medium (medium with ornithine in place of arginine). Variants that have lost this ability are isolated from 77orn by a ‘suicide’ selective technique dependent on the cellular incorporation of [3H]ornithine. These variants, which have reduced levels of CPS-I, or of both CPS-I and OTC, are shown to have developed multiple hormonal requirements; their enzyme deficiencies can be reversed by use of an appropriately supplemented medium. In particular, CPS-I is inducible by dexamethasone and dibutyryl-cyclic-AMP in combination. Cholera toxin can be used instead of cyclic-AMP, but then butyrate is additionally required if the induction is to be maintained in the long term. The use of these agents in excess can depress OTC. Several other hepatomas, and alos explanted foetal rat liver cells, have similar requirements for CPS-I expression. It is argued that multiple hormonal requirements for CPS-I production are normal in liver cells in vitro, and that hormone-independent hepatomas should be regarded as abnormal. The implications of this for the somatic cell genetic investigation of differentiation are briefly discussed.

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