Carrier membrane as a stationary phase for affinity chromatography and kinetic studies of membrane-bound enzymes.
The use of membrane supports as stationary phase, coupled with ligands of choice, allows all kinds of chromatography [Dj. Josić, K. Zeilinger, Y. Lim, M. Raps, W. Hofmann and W. Reutter, J. Chromatogr., 484 (1989) 327] and offers a powerful alternative to both soft gel chromatography and high-performance liquid chromatography. In this work we present affinity membrane chromatography for purification of the enzyme carbonic anhydrase from haemolysates of human erythrocytes. Furthermore, the coupling of the enzymes to the membrane support allows kinetic investigations. As an example, kinetic experiments were carried out by means of carbonic anhydrase coupled to the membrane support using 4-nitrophenyl acetate and 2-chloro-4-nitrophenyl acetate as substrates.
Institut für Molekularbiologie und Biochemie der Freien Universität Berlin, Germany.
H
H
Abou-Rebyeh
F
F
Körber
K
K
Schubert-Rehberg
J
J
Reusch
D
D
Josić
D002256:Carbonic Anhydrases / Q000097:blood*; D002846:Chromatography, Affinity / Q000379:methods; D004800:Enzymes, Immobilized / Q000097:blood*; D004910:Erythrocyte Membrane / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D008567:Membranes, Artificial; D009596:Nitrophenols / Q000378:metabolism; D012995:Solubility; D013379:Substrate Specificity
D004800:Enzymes, Immobilized; D008567:Membranes, Artificial; D009596:Nitrophenols; C071472:2-chloro-4-nitrophenyl acetate; C008642:4-nitrophenyl acetate; D002256:Carbonic Anhydrases