DEAE-cellulose column chromatography

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.]... 

Hoffmann GF, Brendel SU, et al. (1992) Mevalonate kinase assay using DEAE-cellulose column chromatography for first-trimester prenatal diagnosis and complementation analysis in mevalonic aciduria. J Inherit Metab Dis 15 738-746... 

Nagasawa, T., Kiyosawa, I., Kuwahara, K. and Ganguly, N. C. 1973. Fractionation of buffalo milk casein by acrylamide gel electrophoresis and DEAE cellulose column chromatography. J. Dairy Sci. 56, 61-65. 

Ribadeau-Dumas, B., Maubois, J. L., Mocquot, G. and Gamier, J. 1964. A study of casein by DEAE-cellulose column chromatography in urea. Biochim. Biophys. Acta 82, 494-506. 

A mixture of 0.44 g of acidic proteinoid, 0.19 g of basic proteinoid, and 0.31 g of radioactive ATP in 2.0 ml of 0.02 M MgCl2, 0.05 M Tris buffer, is heated in a boiling water bath for 5 min, and then incubated at 37 °C for 24 hrs, after the fractionation of the mixture by DEAE-cellulose column chromatography. Oligonucleotides produced are identified with those of authentic markers 47). Chain length has been determined by the ratio of AMP to adenosine in the alkaline hydrolyzate of the material which was treated by alkaline phosphatase to remove 5 - and 3 -phosphate47). 

The key to success in extensive purification and crystallization of ferredoxin is the use of diethylaminoethyl (DEAE) cellulose column chromatography. Ferredoxin has a marked affinity for DEAE-cellulose, and when a cell-free extract containing ferredoxin is passed through a DEAE-cellulose column, ferredoxin remains as a band at the top while contaminating protein passes through in the effluent. The affinity of ferredoxin for DEAE-cellulose may be explained by its isoelectric point of 3.7 (Lovenberg, Buchanan, and Rabinowitz (65)). 

The reaction mixture (final volume 0.34 ml) contained (in /umol and including compounds added with the enzyme) potassium phosphate, pH 6.3, 102 Tris, pH 7.6, 5 sucrose, 13 GSH, 16 2-mercaptoethanol, 0.7 Na EDTA, 0.5. Methyl donors were added at the specified final concentrations. The various stereoisomeric forms of S-adenosylmethionine were prepared as described by de la Haba el al. (1959) using adenosylmethionine cyclotransferase (E.C. 2.5.1.4) to resolve (/(S)-S-adenosyl-L-methionine to (f )-5-adenosyl-L-methionine. The reaction was started by addition of enzyme dissolved in Tris buffer, pH 7.6, 25 mM, containing Na EDTA, 1 mM and 2-mercaptoethanol, 7 mM, and partially purified from an extract of cabbage leaves by successive ammonium sulfate precipitation (45-60% saturation), gel filtration (Sephadex G-25) and DEAE-cellulose column chromatography. After incubation at 26°C for 1 h, each reaction mixture was diluted with 0.76 ml cold water, and a 1.0 ml aliquot was immediately added to a column of Dowex-50(NHJ) 1 x 3 cm. Unreacted [ C]methionine was removed by washing the column with 25 ml water. The 5-[" C]methylmethionine sulfonium salt was then eluted with 8 ml 0.6 N NH OH. Radioactivity was determined by scintillation spectrophotometry (Mudd et al., 1965). A blank of 540 cpm has been subtracted from each value. The low activity with (R)-5-adenosyl-L-methionine may have been due to the presence of a small residual amount of (5)-5-adenosyl-L-methionine (de la Haba eta/., 1959). 

The enzyme can be purified from the microsomes of bovine vesicular gland through DEAE-cellulose column chromatography, isoelectrofocusing and Sepharose column chromatography. The purified enzyme shows an apparent homogeneity upon poly-... 

The DEAE-cellulose column chromatography of pectic substances was carried out by the method of Plat et al (1991) with a slight modification. Pectic substances (in 50 mM sodium phosphate buffer, pH 6.3) were added to a DEAE cellulose column (2 x 40 cm) equilibrated with the same phosphate buffer. The solution was then eluted with a 50 mM sodium phosphate buffer (pH 6.3) with a flow rate of 30 mL/h. The fraction (2 mL) was collected and monitored by the carbazole colorimetric method and expressed as the absorbance at 525 nm. 

Purified by DEAE-cellulose column chromatography Amberlite XAD-2 Pure C-2801X, sodium salt... 

Turner et al. (305) isolated the deacetoxycephalosporin C oxygenase from both C. acremonium and S. clavuligerus by DEAE-cellulose column chromatography. They concluded that the enzymes were dioxygenases as they both required a-ketoglutarate, ferrous ions, ascorbate and dithio-threitol for full activity. The C. acremonium enzyme was markedly activated by preincubation with ferrous ions. Both enzymes where inhibited by high concentrations of nucleotides. The pH optimum of the S. clavuligerus enzyme was about pH 7.0, whereas that of C. acremonium was about pH 6.0. Neither enzyme would accept the deacetoxycephalosporins (302)—(307) as substrates. 

DEAE-cellulose column chromatography has contributed much to the advancement of purification and separation of multiple forms of glucuronidase (Moore and Lee, 1960). 

Abbreviations LWAF - long-wavelength absorbing form, DEAEcc - DEAE cellulose column chromatography. 

Wi ckowski S, Droba M (1978) Heterogeneity of the thylakoid membrane fractions derived from spinach chloroplasts by the action of Triton X-100 in low salt medium. II. Isolation of / -carotene by the DEAE cellulose column chromatography. Plant Sc.Lett. 13, 397-404. 

---