DEAE-cellulose Sephadex

Azotobacter chroococcum French pressure cell DEAE-cellulose Sephadex... 

Rhizobium japonicum bacteroids French pressure cell Polypropylene glycol DEAE-cellulose Sephadex G-200... 

Abras precatorius. Purified by successive chromatography on DEAE-Sephadex A-50, carboxymethylcellulose, and DEAE-cellulose. [Wei et al. J Biol Chem 249 3061 1974.]... 

Purified by Sephadex G-200 filtration and DEAE-cellulose column chromatography. Hexosaminidase A was further purified by DEAE-cellulose column chromatography, followed by an ECTEOLA-cellulose column, Sephadex-200 filtration, electrofocusing and Sephadex G-200 filtration. Hexosaminidase B was purified by a CM-cellulose column, electrofocusing and Sephadex G-200 filtration. [Srivastava et al. 7 Biol Chem 249 2034 1974.]... 

Dibydropteridine reductase (from sbeep liver) [9074-11-7] Mr 52,000 [EC 1.6.99.7]. Purified by fractionation with ammonium sulfate, dialysed versus tris buffer, adsorbed and eluted from hydroxylapatite gel. Then run through a DEAE-cellulose column and also subjected to Sephadex G-lOO filtration. [Craine et al. J Biol Chem 247 6082 1972.]... 

Reverse transcriptase (from avian or murine RNA tumour viruses) [9068-38-6] [EC 2.7.7.49]. Purified by solubilising the virus with non-ionic detergent. Lysed virions were adsorbed on DEAE-cellulose or DEAE-Sephadex columns and the enzyme eluted with a salt gradient, then chromatographed on a phosphocellulose column and enzyme activity eluted in a salt gradient. Purified from other viral proteins by affinity chromatography on a pyran-Sepharose column. [Verna Biochim Biophys Acta 473 1 7977 Smith Methods Enzymol 65 560 1980 see commercial catalogues for other transcriptases.]... 

Thrombin (from bovine blood plasma) [9002-04-4] Mj 32,600 [EC 3.4.4.13]. Purified by chromatography on a DEAE-cellulose column, while eluting with O.IM NaCl, pH 7.0, followed by chromatography on Sephadex G-200. Final preparation was free from plasminogen and plasmin. [Yin and Wessler J Biol Chem 243 112 796S.]... 

Dried shrimp was ground, defatted with benzene, and then extracted with cold water. The luciferase extracted was purified first by a batch adsorption onto DEAE cellulose (elution with 0.4 M NaCl), followed by gel filtration on a column of Sephadex G-150, anion-exchange chromatography on a column of DEAE-cellulose (gradient elution 0.05-0.5 M NaCl), and gel filtration on a column of Ultrogel AcA 34. The specific activity of the purified luciferase was 1.7 x 1015 photons s 1 mg-1, and the yield in terms of luciferase activity was about 28%. 

Purification of photoprotein. The dialyzed photoprotein solution was centrifuged to remove precipitates, and then subjected to fractional precipitation by ammonium sulfate, taking a fraction precipitated between 30% and 50% saturation. The protein precipitate was dissolved in 50 ml of 10 mM sodium phosphate, pH 6.0, containing 0.1 mM oxine ( pH 6.0 buffer ), dialyzed against the same buffer, and the dialyzed solution was adsorbed on a column of DEAE-cellulose (2.5 x 13 cm) prepared with the pH 6.0 buffer. The elution was done by a stepwise increase of NaCl concentration. The photoprotein was eluted at 0.2-0.25 M NaCl and a cloudy substance (cofactor 1) was eluted at about 0.5 M NaCl. The photoprotein fraction was further purified on a column of Sephadex G-200 or Ultrogel AcA 34 (1.6 x 80 cm) using the pH 6.0 buffer that contained 0.5 M NaCl. 

Step 3. The photoprotein fractions are combined and the NaCl concentration of the solution is reduced to 50 mM by gel filtration through a column of Sephadex G-25. Then, the solution is poured onto a column of DEAE cellulose. The photoprotein adsorbed on the column is eluted by a linear increase of NaCl concentration from 50 mM to 0.4 M in the same pH 6.7 buffer. Active fractions are combined and concentrated by ammonium sulfate precipitation. 

Macrochromatographlc Methods. These procedures use cation exchangers, such asAmberllte-IRC-50, carboxymethyl-Cellulose (CMC) and carboxymethyl-Sephadex (CMS), and the anion exchangers diethylamlnoethyl-(DEAE)-Cellulose and DEAE-Sephadex. 

Chromatography of hemoglobins on columns of DEAE-Cellulose (DE-52 mlcrogranular, preswollen Whatman) often results In excellent separations of many variants, because the hemoglobins are eluted as sharp, narrow zones generally widely separated from each other The hemoglobin zones are eluted from the DE-52 columns at a distinctly higher pH value than from a similar DEAE-Sephadex column ... 

In the purification of pectinesterase from the fruits of Citrus nat-sudaidai,61 fractional salting-out with ammonium sulfate was followed by chromatography on a column of DEAE-cellulose and by separation of the active fraction on Sephadex G-100. A preparation (purified solution) having a specific activity 460-fold greater than that of the original extract was obtained. Its homogeneity was checked by disc electrophoresis, and its amino acid content was determined and fundamental, kinetic data were obtained. 

Macmillan and coworkers51 105 106 purified pectinesterase produced by Clostridium multifermentans by using practically all of the available methods and materials (calcium phosphate gel, DEAE-cellulose, DEAE-, QEAE-, CM-, and SE-Sephadex, Sephadexes G-75, G-100, G-150, and G-200, Sepharose 4B, and zonal centrifugation). However, they could not separate pectinesterase from exo-pectate lyase, and, hence, they postulated that either (a) a complex of the two enzymes having an apparent molecular weight of 400,000 exists, or (b) the two enzymes are identical in their molecular species. On the basis of the mode of action of this pectinesterase in comparison with that of those from tomatoes and from Fusarium ox-ysporum, the existence of a complex of pectinesterase and exopectate lyase in Clostridium multifermentans appears to be the more probable. 

Adsorption on calcium pectate and calcium phosphate, and chromatography on DEAE-cellulose, were used for the purification of pectin lyase from Aspergillus fonsecaeus.253 Two forms of pectin lyase (having pH optima at 7.3 and 8.3) were isolated266 from the culture filtrate of Sclerotinia fructigena by chromatography on Seph-adex G-75 and CM-Sephadex C-50. 

Xylanase was obtained from filtered fermentation broths of Chainia NCL-82-5-1 (6). It was precipitated by the addition of three volumes of 0-4°C ethanol, separated from the supernatant by centrifugation, and lyophilized. Four column chromatographic steps, DEAE-cellulose anion exchange, Fractogel TSK DEAE-650S anion exchange, Sephadex G-50-50 gel permeation, and CM-Trisacryl cation exchange, were used for pu ication. 

In various runs, specific activity went from an average of 2.75 lU/mg protein (based on the Bio-Rad protein assay with bovine serum albumin standard) for Ae centrifuged crude preparation after alcohol precipitation, to 8 lU/mg after DEAE-cellulose chromatography, to 100 lU/mg after DEAE-Fractogel chromatography, to 1000 RJ/mg after Sephadex G-50 chromatography, and to 3200 lU/mg after CM-Trisacryl chromatography. 

Column Chromatography of Crude Toxin. The WSAP obtained from culture 350F was retained in the crude state for assay. The 266.2 mg of WSAP obtained from 350G was treated on successive columns of silicic acid, DEAE cellulose, and Sephadex G-15 and yielded a single semipurified toxic product (GT-4) of 23.1 mg or 9% of the starting crude extract (Table I). 

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