THE ROLE OF PROTEIN CHARGE IN PROTEIN LIPID INTERACTIONS - PH-DEPENDENT CHANGES OF THE ELECTROPHORETIC MOBILITY OF LIPOSOMES THROUGH ADSORPTION OF WATER-SOLUBLE, GLOBULAR-PROTEINS

The role of electrostatics in the adsorption process of proteins to pr eformed negatively-charged (phosphatidylcholine/phosphatidylglycerol) and neutral (phosphatidylcholine) liposomes was studied. The interacti on was monitored at low ionic strength for a set of model proteins as a function of pH. The adsorption behavior of trypsin inhibitor (pI = 4 .6), myoglobin (pI = 7.4), ribonuclease (pI = 9.6), and lysozyme (pI = 10.7) with preformed liposomes was investigated, along with changes i n the electrophoretic mobility of liposomes through the adsorption of charged proteins. Mean protein charge was determined by acid/base titr ation. Significant adsorption of the proteins to negatively-charged li posomes was only found at pH values where the number of positive charg e moieties exceeds the number of negative charge moieties on the prote in by at least three charge units. Negligible adsorption to liposomes composed of zwitterionic lipids was observed in the pH range tested (4 -9). The absolute value of the electrophoretic mobilities of negativel y-charged, empty liposomes decreased after adsorption of positively-ch arged proteins. With increasing protein to phospholipid ratio, the dro p in the electrophoretic mobility leveled off and reached a plateau; p rotein adsorption profiles showed a similar shape. Analysis of the dat a demonstrated that neutralization of the liposome charge due to the a dsorption of the positively-charged proteins is the controlling factor in their adsorption. The plateau level reached depended on the type o f protein and the pH of the incubation medium. This pH dependency coul d be ascribed to the mean positive charge of the protein. The effectiv e charge of myoglobin, ribonuclease, and lysozyme (defined as the numb er of phosphatidylglycerol groups neutralized by one adsorbed protein molecule) was calculated from the charge differences between empty lip osomes and protein-coated liposomes using the Gouy-Chapman theory. For lysozyme and myoglobin, an excellent correlation was found between th e effective charge and the mean protein charge.