Complexation of Papain with Strong Polyanions and Enzymatic Activities of the Resulting Complexes

Abstract

The complexation of papain (EC.3.4.22.2) with potassium polyvinyl alcohol) sulfate (KPVS) and sodium poly(styrene sulfonate) (NaPSS) was studied at different pH levels by means of colloid titration. The enzymatic activities of the resulting complexes were also studied as a function of pH and temperature, using Nα-benzoyl-DL-arginine-p-nitroanilide as the substrate. The moles (M ^s)of the sulfate or sulfonate groups in the polyelectrolytes that took part in the complexation with 1 g papain varied depending on pH due to a pH-induced change in the protonation of the protein basic groups: 11 amino (including one N-terminal), 2 imidazolyl, and 12 guanidyl groups. However, the curves of M ^s vs pH were independent of neither the species of the polyelectrolytes nor the molecular weight of NaPSS. On the other hand, the pH-activity curve of papain was shifted toward the alkaline pH range upon complexation, which occurred only in a region of pH < 5.5, without an accordant shift of the optimum pH (near 6.5 for both native and complexed enzymes). A loss in the enzyme activity due to complexation was observed of the magnitude of that of NaPSS-50 (MW = 1,260,000) > KPVS≅ NaPSS-7 (MW = 68,000). The mechanism of formation and structure of the protein/polyelectrolyte complexes consisting of papain and KPVS or NaPSS are discussed on the basis of these results, particularly in terms of the effect of the hydrophobicity of polyion chains on the structure of the complexes.