title: Purification of human erythrocyte adenosine deaminase by affinity column chromatography.
abstract: Adenosine deaminase (adenosine aminohydrolase EC 3.5.4.4) has been purified 468,000-fold from pooled human erythrocytes. The procedure developed was used to isolate the enzyme from up to 23 liters of packed erythrocytes at one time. An easily prepared affinity column bed material employing adenosine as the ligand was used as the final step in the purification. During elution from the affinity column there was approximately a 3:1 partition of adenosine deaminase between gel bed and column buffer. There was no apparent difference in the partitioning of unresolved or partially resolved preparations of the electrophoretically different forms of the enzyme on the affinity column. Gel filtration and electrophoresis on polyacrylamide gels of increasing concentration revealed no differences in the Mr of these electrophoretically different forms. The four bands resolved by electrophoresis of the different forms on polyacrylamide gels under nondenaturing conditions yielded a single band when electrophoresis was carried out in the presence of sodium dodecyl sulfate and 2-mercaptoethanol. Partially resolved preparations of the different electrophoretic forms of adenosine deaminase also gave rise to a single band of the same mobility when electrophoresed on polyacrylamide gels  under these conditions. The band had the mobility of a protein of Mr of 36,000. This Mr is approximately the same as estimated for the nondenatured enzyme.
mesh_terms: D000243:Adenosine Deaminase / Q000097:blood* / Q000302:isolation & purification; D002846:Chromatography, Affinity; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D008970:Molecular Weight; D009700:Nucleoside Deaminases / Q000097:blood*
chemical_list: D009700:Nucleoside Deaminases; D000243:Adenosine Deaminase