Solid-phase enzyme modification via affinity chromatography

Abstract

In the present study antileukemic enzyme L-asparaginase (ASNase) and catalase (as a model enzyme) were modified in solid-phase with activated polyethylene glycol (PEG ) by using ligand-immobilized affinity column systems L-asparagine- 2 Sepharose CL-4B and Procion red-Sepharose CL-4B, respectively. Studies on change of specific activity with modification time showed negligible differences between batches of modified catalase. Modification of ASNase for 1 h resulted in 50.2% recovery of the specific activity and the attachment of 69 molecules of PEG per molecule of ASNase forming ‘PEGylated 2 ASNase’. Sequential modification of ASNase by activated PEG and heparin resulted in coupling of about nine molecules of heparin per molecule of PEGylated ASNase. Intravenous (i.v.) administration of PEG -modified ASNase showed prolonged 2 presence in the blood circulation and no adverse effects or symptoms of anaphylaxis were observed in presensitized mice.