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 PROTEIN CHARGE IN PROTEIN LIPID INTERACTIONS - PH-DEPENDENT CHANGES OF THE ELECTROPHORETIC MOBILITY OF LIPOSOMES THROUGH ADSORPTION OF WATER-SOLUBLE, GLOBULAR-PROTEINS
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DOI：
10.1021/bi00068a023
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发表时间：
1993-05-04
期刊：
影响因子：
2.9
通讯作者：
CROMMELIN, DJA
中科院分区：
文献类型：
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作者：
BERGERS, JJ;VINGERHOEDS, MH;CROMMELIN, DJA
研究了静电在蛋白质吸附于预成型的带负电荷（磷脂酰胆碱/磷脂酰甘油）和中性（磷脂酰胆碱）脂质体过程中的作用。在低离子强度下监测一组模型蛋白质的相互作用作为pH值的函数。胰蛋白酶抑制剂（pI = 4.6），肌红蛋白（pI = 7.4），核糖核酸酶（pI = 9.6），溶菌酶（pI = 10.7）与预制脂质体的吸附行为进行了研究，沿着通过带电蛋白质的吸附在脂质体的电泳迁移率的变化。通过酸/碱滴定法测定平均蛋白质电荷。仅在正电荷部分的数量超过蛋白质上的负电荷部分的数量至少三个电荷单位的pH值下发现蛋白质对带负电荷的脂质体的显著吸附。在测试的pH范围内（4-9），观察到对由两性离子脂质组成的脂质体的吸附可忽略不计。带负电荷的空脂质体的电泳迁移率的绝对值下降后，吸附带正电荷的蛋白质。随着蛋白质与磷脂比例的增加，电泳迁移率的下降趋于平稳并达到平台;蛋白质吸附曲线显示出类似的形状。数据分析表明，由于带正电荷的蛋白质的吸附，脂质体电荷的中和是其吸附的控制因素。达到的平台水平取决于蛋白质的类型和孵育介质的pH值。这种pH依赖性可归因于蛋白质的平均正电荷。肌红蛋白、核糖核酸酶和溶菌酶的有效电荷（定义为被一个吸附的蛋白质分子中和的磷脂酰甘油基团的数量）使用Gouy-Chapman理论从空脂质体和蛋白质包被的脂质体之间的电荷差计算。对于溶菌酶和肌红蛋白，一个很好的相关性之间的有效电荷和平均蛋白质电荷。
The role of electrostatics in the adsorption process of proteins to preformed negatively-charged (phosphatidylcholine/phosphatidylglycerol) and neutral (phosphatidylcholine) liposomes was studied. The interaction 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 in the electrophoretic mobility of liposomes through the adsorption of charged proteins. Mean protein charge was determined by acid/base titration. Significant adsorption of the proteins to negatively-charged liposomes was only found at pH values where the number of positive charge moieties exceeds the number of negative charge moieties on the protein 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 negatively-charged, empty liposomes decreased after adsorption of positively-charged proteins. With increasing protein to phospholipid ratio, the drop in the electrophoretic mobility leveled off and reached a plateau; protein adsorption profiles showed a similar shape. Analysis of the data demonstrated that neutralization of the liposome charge due to the adsorption of the positively-charged proteins is the controlling factor in their adsorption. The plateau level reached depended on the type of protein and the pH of the incubation medium. This pH dependency could be ascribed to the mean positive charge of the protein. The effective charge of myoglobin, ribonuclease, and lysozyme (defined as the number of phosphatidylglycerol groups neutralized by one adsorbed protein molecule) was calculated from the charge differences between empty liposomes and protein-coated liposomes using the Gouy-Chapman theory. For lysozyme and myoglobin, an excellent correlation was found between the effective charge and the mean protein charge.