Effects of surface charge distribution of proteins in their complexation with polyelectrolytes in an aqueous salt-free system

Complexation between protein and polyelectrolyte was studied by a combinati on of three main experimental techniques: turbidimetric titration, quasi-el astic light scattering (QELS), and static light scattering (SLS). Proteins of two sorts, lysozyme (Lyz) and ribonuclease (RNase), were chosen by consi dering the following characteristics: (i) Both proteins have the same numbe r (19) of basic groups; (ii) their distribution is almost homogeneous on Ly z but not on RNase; (iii) there is little difference in the molar mass betw een both proteins. Potassium poly(vinyl alcohol) sulfate (KPVS) with differ ent molecular weights ((M) over bar(PE)) and various degrees of esterificat ion (D-e) was used as the polyelectrolyte. We employed a salt-free aqueous medium and adjusted it to pH 2, the level of which forces to completion the protonation of all of the basic groups. As the titration of proteins with KPVS proceeded, the absorbance (A) as an indication of turbidity increased linearly and then rapidly at a certain titrant volume, referring to the end point of titration. The slope of the linear plots of A vs titrant volume f or Lyz was Little dependent on (M) over bar(PE) and D-e. In the case of RNa se, however, the slope increased with decreasing D-e and with increasing (M ) over bar(PE). From the studies of the hydrodynamic radius by QELS as well as the molecular weight and radius gyration by SLS, it was found that the (M) over bar(PE) and D-e effects observed in the titration curve correspond to the changes of both size and mass of aggregated "intrapolymer" complexe s formed during titration. Thus, we estimated the degree of aggregation (al pha) through dividing the mass from SLS by the calculated mass of an intrap olymer complex. This showed that alpha decreases with increasing D-e in bot h protein systems, while increasing (M) over bar(PE) decreases alpha in the Lyz system but does not change it in the RNase system. These results were discussed in detail by considering the polarizability of the intrapolymer c omplex, the level of which is affected by the complementarity of the spacin g between charges on the protein to the uniform spacing between polymer cha rges.