Affinophoresis of trypsins.
We devised a new separation technique for the protein, "affinophoresis," which is based on its specific affinity and utilizes electrophoresis. This technique requires a carrier macromolecule, "affinophore," which contains both an affinity ligand for a certain protein and many charges, either positive or negative, in order to migrate rapidly in an electric field. When a mixture of proteins is electrophoresed in the presence of the affinophore, the protein having an affinity with the ligand will form a complex with the affinophore. This results in a change in the apparent electrophoretic mobility. If the protein is sufficiently accelerated, we can separate it from other materials. A cationic affinophore for trypsin was prepared. Soluble dextran MW approximately 10,000) was coupled with a DEAE-group and m-aminobenzamidine, a competitive inhibitor of trypsins. Electrophoresis of trypsins from several origins on agarose gel plates in the presence of the affinophore showed that affinophoresis actually occurred. The electrophoretic mobilities of trypsins increased towards the cathode, the same direction as the affinophore movement. The presence of leupeptin and treatment of the trypsins with TLCK suppressed the effect of the affinophore. Streptomyces griseus trypsin, contained in Pronase, was easily separated and detected. This procedure is distinct from affinity chromatography and so-called affinity electrophoresis in that the support of the affinity ligand moves, and has advantages especially for analytical purposes: for example, the detection of specific molecules regardless of their isoelectric points.
D000345:Affinity Labels; D000614:Aminocaproates; D000818:Animals; D002417:Cattle; D002846:Chromatography, Affinity / Q000379:methods; D003911:Dextrans; D004586:Electrophoresis / Q000379:methods; D004983:Ethanolamines; D007202:Indicators and Reagents; D014357:Trypsin / Q000302:isolation & purification*
D000345:Affinity Labels; D000614:Aminocaproates; D003911:Dextrans; D004983:Ethanolamines; D007202:Indicators and Reagents; C018802:6-acetylaminocaproic acid; D014357:Trypsin; C007369:2-diethylaminoethanol