New strategies for the screening of a large number of immobilized dyes for the purification of enzymes. Application to the purification of enzymes from human haemolysate.
A method is presented for screening immobilized dyes applicable to the purification of enzymes from haemolysate (haemolysate can be considered as a nearly pure solution of haemoglobin containing only marginal amounts of enzymes). Haemolysate is loaded on immobilized dye mini-columns until haemoglobin and the studied enzymes are found in the column eluate at the same concentrations as those present in the haemolysate. Such a frontal mode of screening allows those dyes to be selected which, displaying a higher affinity for the enzyme of interest than for haemoglobin, can be used to displace the unwanted protein (haemoglobin) from the column by the enzyme of interest (present at a much lower concentration).
Kroviarski
Y
Y
INSERM U160, Hôpital Beaujon, Clichy, France.
Cochet
S
S
Vadon
C
C
Truskolaski
A
A
Boivin
P
P
Bertrand
O
O
D004396:Coloring Agents; D004798:Enzymes / Q000097:blood* / Q000302:isolation & purification; D004912:Erythrocytes / Q000201:enzymology; D005956:Glucose-6-Phosphate Isomerase / Q000097:blood / Q000302:isolation & purification; D006461:Hemolysis; D006801:Humans; D009249:NADP / Q000378:metabolism; D010734:Phosphogluconate Dehydrogenase / Q000097:blood / Q000302:isolation & purification; D012685:Sepharose
D004396:Coloring Agents; D004798:Enzymes; D009249:NADP; D012685:Sepharose; D010734:Phosphogluconate Dehydrogenase; D005956:Glucose-6-Phosphate Isomerase