scholarly journals The rapid purification of 3-hydroxybutyrate dehydrogenase and malate dehydrogenase on triazine dye affinity matrices

1982 ◽  
Vol 203 (3) ◽  
pp. 699-705 ◽  
Author(s):  
M D Scawen ◽  
J Darbyshire ◽  
M J Harvey ◽  
T Atkinson
Keyword(s):  
Affinity Chromatography ◽  
Malate Dehydrogenase ◽  
Large Scale ◽  
Gel Filtration ◽  
Rapid Purification ◽  
Dye Affinity Chromatography ◽  
Triazine Dye

3-Hydroxybutyrate dehydrogenase (EC 1.1.1.30) and malate dehydrogenase (EC 1.1.1.37) were purified to homogeneity on a large scale involving only two sequential affinity-chromatography steps on two triazine dye-Sepharose matrices. Recoveries of both enzymes were in excess of 60%. Malate dehydrogenase could also be purified by a combination of triazine dye affinity chromatography and gel filtration on Ultrogel AcA-44, but this offered no significant advantage over the purely affinity procedure.

A Combination of Cation Exchange and Ligand-Affinity Chromatography for Purification of Two Molecular Species of the Folate Binding Protein in Human Milk, One Equipped with a Hydrophobic Glycosyl Phosphatidylinositol Tail: Characterization of Hydrophobicity and Electrical Charge

2002 ◽  
Vol 22 (3-4) ◽  
pp. 443-454 ◽  
Author(s):  
Jan Holm ◽  
Steen Ingemann Hansen ◽  
Mimi Høier-Madsen
Keyword(s):  
Human Milk ◽  
Affinity Chromatography ◽  
Cation Exchange ◽  
Large Scale ◽  
Gel Filtration ◽  
Molecular Size ◽  
Exchange Chromatography ◽  
Amino Acid Sequences ◽  
Affinity Column ◽  
Folate Binding Protein

Cation exchange chromatography combined with ligand (methotrexate) affinity chromatography on a column desorbed with a pH-gradient was used for separation and large scale purification of two folate binding proteins in human milk. One of the proteins, which had a molecular size of 27 kDa on gel filtration and eluted from the affinity column at pH 5–6 was a cleavage product of a 100 kDa protein eluted at pH 3–4 as evidenced by identical N-terminal amino acid sequences and a reduction in the molecular size of the latter protein to 27 kDa after cleavage of its hydrophobic glycosylphosphatidyl-inositol tail that inserts into Triton X-100 micelles. Chromatofocusing showed that both proteins possessed multiple isoelectric points within the pH range 7–9. The 100 kDa protein exhibited a high affinity to hydrophobic interaction chromatographic gels, whereas this was only the case with unliganded forms of the 27 kDa protein indicative of a decrease in the hydrophobicity of the protein after ligand binding.

Triazine-dye affinity chromatography

1981 ◽  
Vol 9 (4) ◽  
pp. 290-293 ◽  
Author(s):  
TONY ATKINSON ◽  
PETER M. HAMMOND ◽  
ROY D. HARTWELL ◽  
PETER HUGHES ◽  
MICHAEL D. SCAWEN ◽  
...  
Keyword(s):  
Affinity Chromatography ◽  
Dye Affinity Chromatography ◽  
Triazine Dye

Purification of the Antihemophilic Factor by Gel Filtration on Agarose

1969 ◽  
Vol 22 (03) ◽  
pp. 577-583 ◽  
Author(s):  
M.M.P Paulssen ◽  
A.C.M.G.B Wouterlood ◽  
H.L.M.A Scheffers
Keyword(s):  
Factor Viii ◽  
Plasma Proteins ◽  
Large Scale ◽  
Gel Filtration ◽  
Large Scale Preparation ◽  
Scale Preparation ◽  
Antihemophilic Factor

SummaryFactor VIII can be isolated from plasma proteins, including fibrinogen by chromatography on agarose. The best results were obtained with Sepharose 6B. Large scale preparation is also possible when cryoprecipitate is separated by chromatography. In most fractions containing factor VIII a turbidity is observed which may be due to the presence of chylomicrons.The purified factor VIII was active in vivo as well as in vitro.

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