Affinity chromatography in the separation of human alpha 1-antitrypsin (alpha 1-AT) and antithrombin-III (AT-III).
The two antiproteases alpha 1-antitrypsin (alpha 1-AT) and antithrombin-III (AT-III) have been purified simultaneously from human plasma. Purification procedure consisted of gel filtration on Sephadex G-200 after initial processing of plasma, followed by ion exchange chromatography on DEAE-Sephadex A50 and DEAE-Cellulose, at a pH of 9.0 and pH 8.3 respectively. The two proteins could not be separated by any of these procedures including a lower pH (7.4) in ion exchange chromatography. Affinity chromatography on heparin-Sepharose separated the proteins since alpha 1-AT did not bind to the matrix. Alpha 1-AT unbound to the heparin-Sepharose was subsequently purified through con A-Sepharose affinity column. The final yield of both the proteins was about 20%. The molecular weight estimated on SDS electrophoresis for AT-III and alpha 1-AT was 63,000 and 50,000, respectively.
D000990:Antithrombin III / Q000302:isolation & purification*; D002846:Chromatography, Affinity / Q000379:methods; D003208:Concanavalin A; D006493:Heparin; D006801:Humans; D012685:Sepharose; D000515:alpha 1-Antitrypsin / Q000302:isolation & purification*
D000515:alpha 1-Antitrypsin; D003208:Concanavalin A; D000990:Antithrombin III; D006493:Heparin; D012685:Sepharose