Search results for "Affinity chromatography"
showing 10 items of 82 documents
Association of a polyuridylate-specific endoribonuclease with small nuclear ribonucleo-proteins which had been isolated by affinity chromatography us…
1983
Immunoglobulins, containing antibodies against U1-snRNP, have been prepared from a patient with systemic lupus erythematosus. After coupling these antibodies to a Sepharose matrix, U-snRNPs have been isolated and purified from rat liver nuclei by use of immunoaffinity chromatography. The resulting RNPs had the typical protein pattern of U-sn RNPs and a sedimentation coefficient of 12 S. The U-snRNP preparation was associated with an endoribonuclease which required Mg2+ for optimal activity. The enzyme, with an pH optimum of 6.2, degraded only poly(U). Other single-stranded polyribo- and polydeoxyribonucleotides, tRNA, as well as double-stranded RNA and DNA were not digested. The products of…
Characterization of rat glutathione transferases in olfactory epithelium and mucus
2019
International audience; The olfactory epithelium is continuously exposed to exogenous chemicals, including odorants. During the past decade, the enzymes surrounding the olfactory receptors have been shown to make an important contribution to the process of olfaction. Mammalian xenobiotic metabolizing enzymes, such as cytochrome P450, esterases and glutathione transferases (GSTs), have been shown to participate in odorant clearance from the olfactory receptor environment, consequently contributing to the maintenance of sensitivity toward odorants. GSTs have previously been shown to be involved in numerous physiological processes, including detoxification, steroid hormone biosynthesis, and am…
The major isozyme of rat cardiac glutathione transferases. Its correspondence to hepatic transferase X.
1986
1. A major isozyme of rat heart glutathione transferase was purified to homogeneity by Sephadex G-200 gel filtration, ammonium sulfate precipitation, CM-cellulose chromatography and affinity chromatography on S-hexylglutathione-linked Sepharose 6B. 2. The purified isozyme was a dimer with an apparent relative molecular mass of 50000 composed of two Yb-size subunits (Mr= 26 500). The isozyme is immunologically related to rat liver glutathione transferase X and 3–3, especially closely to transferase X, and no immunological cross-reactivity with subunits 1 and 2 of hepatic glutathione transferases was observed. The isoelectric point (pI = 6.9) of the isozyme was identical with and the substrat…
Development of HPLC methods for the purification and analysis of plasma membrane glycoproteins.
1990
High resolution HPLC techniques such as affinity chromatography (AC), ion exchange chromatography (IEC), and size exclusion chromatography (SEC) were used successfully for separations of hydrophobic plasma membrane glycoproteins. We have tested a lot of commercially available columns for IEC and SEC and performed the purification of the crude plasma membrane extract with the most suitable columns. By using immobilized ligands with different specificities and sequential affinity chromatography, it is possible to obtain a preliminary structural characterization of the interesting carbohydrate residues of membrane glycoproteins.
The use of trypsin to solubilize wall proteins from Candida albicans led to the identification of chitinase 2 as an enzyme covalently linked to the y…
2002
The use of trypsin to break proteins covalently linked to the yeast walls of Candida albicans released approx. 50% of the proteins, but also glucose and N-acetylglucosamine. Analysis by affinity chromatography indicated that glucose and/or N-acetylglucosamine formed part of the same supramolecular complexes with mannoproteins. These complexes would represent a new type of cell wall structuration in which beta-1,6 glucan and chitin are linked to proteins. An internal peptide from a 50-kDa protein released by trypsin was sequenced, showing 100% identity with chitinase 2 protein and 92% with chitinase 3. The electrophoretic mobility of the chitinase 2 protein was changed by treatment with Endo…
A Newly-Detected Reductase fromRauvolfiaCloses a Gap in the Biosynthesis of the Antiarrhythmic Alkaloid Ajmaline
2002
A new enzyme, 1,2-dihydrovomilenine reductase (E.C. 1.3.1), has been detected in Rauvolfia cell suspension cultures. The enzyme specifically converts 2beta( R)-1,2-dihydrovomilenine through an NADPH-dependent reaction into 17-O-acetylnorajmaline, a close biosynthetic precursor of the antiarrhythmic alkaloid ajmaline from Rauvolfia. A five-step purification procedure using SOURCE 30Q chromatography, hydroxyapatite chromatography, 2',5'-ADP Sepharose 4B affinity chromatography and ion exchange chromatography on DEAE Sepharose and Mono Q delivered an approximately 200-fold enriched enzyme in a yield of approximately 6%. SDS-PAGE showed an M r for the enzyme of approximately 48 kDa. Optimum pH …
A sensitive monoclonal antibody-based enzyme-linked immunosorbent assay to quantify Parietaria judaica major allergens, Par j 1 and Par j 2
2006
Summary Background Parietaria pollen is one of the most important causes of pollinosis in Mediterranean countries. Parietaria judaica pollen extract presents two major allergens, Par j 1 and Par j 2, that belong to the lipid transfer protein family. Objective To develop an ELISA for quantification of both major allergens of P. judaica pollen extracts, and to assert correlation of these allergens content with the allergenic activity of extracts. Methods Natural Par j 1–Par j 2 allergens were purified by gel filtration, ion exchange, and affinity chromatography and identified by mass spectrometry. Rabbit antisera were obtained using this protein preparation as antigen and used for immunoaffin…
Sugar specific cellular lectins of Phallusia mamillata hemocytes: Purification, characterization and evidence for cell surface localization
1989
Cellular lectins (CLs) of Phallusia mamillata were demonstrated in protein preparations obtained by salt fractionation from hemocytes sonicated in a suitable medium. Since the lectins from the precipitated fraction bind sugars containing D-galactosyl groups, they were purified by affinity chromatography on Sepharose. SDS-PAGE under reducing conditions showed that CLs are formed of two components of apparent MWs approximately 36,900 and 35,090 and thus differ from serum lectins (SLs) (MW about 62,200). The "shrinkage" observed when SLs were examined under nonreducing conditions suggest the presence of intrachain disulphide bonds which can affect the molecular structure of the SLs. CL-SL diff…
Isolation of endothelin A receptor from bovine lungs.
1995
We isolated endothelin receptor A (ET A ) from bovine lungs in a single-step purification procedure using antibodies raised against synthetic peptides that correspond to extra- and intracellular domains of the rat bradykinin receptor. Two receptor species of 55 and 35 kDa were isolated and subjected to N-terminal microsequencing. The difference between the observed and expected molecular weight species suggests that bovine ET A receptor is glycosylated.
Biotin-Labelled and Photoactivatable Aldosterone and Progesterone Derivatives as Ligands for Affinity Chromatography, Fluorescence Immunoassays and P…
1996
New derivatives of progesterone and aldosterone were synthesized and functionally tested with commercially available antibodies. The covalent labelling of antibodies specific for aldosterone and progesterone was detected by SDS/PAGE analysis and subsequent autoradiography after using 3-(O-carboxymethyl)-oximino-(3-[125I]iodo-4-azidosalicylamidobu tylamine) derivatives of aldosterone and progesterone, respectively, as photoactivatable radioligands. Labelling was not observed in the presence of an excess of the unlabelled steroid. Aldosterone was labelled with biotin and used as a tracer in a time-resolved fluorescence immunoassay. The nonradioactive tracer is highly selective for its antibod…