Affinity Chromatography of Trypsin and Related Enzymes II. An Affinity Adsorbent Containing Glycylglycyl-L-arginine1

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Abstract
An affinity adsorbent for trypsin [EC 3.4.21.4] (GGA Sepharose) was prepared. Glycylglycyl-L-arginine (GGA) was synthesized by a simple procedure and was immobilized on agarose gel. This adsorbent proved to have essentially the same characteristics as AP Sepharose, which is an affinity adsorbent containing tryptic peptides of protamine (1). GGA Sepharose was specific for native trypsin and had a stronger affinity at lower pH's (6.5) than at the optimum pH of trypsin action (8.2). It also proved to be suitable for analytical experiments because of its relatively weak affinity. By comparison of the elution profiles of trypsin from GGA Sepharose under various conditions, the nature of the interaction of trypsin with the adsorbent could be studied. It was found that α- and β-trypsin could be distinguished. In the presence of arginine and N-substituted arginines, the elution of trypsin was accelerated. From the extents of the accelerating effects, the affinities of these compounds could be compared.