Welling G W, van der Zee R, Welling-Wester S
J Chromatogr. 1987 Jul 17;418:223-43. doi: 10.1016/0378-4347(87)80010-5.
Biological membranes have as a major function the compartmentation of biological processes in cells and organelles. They consist of a bilayer of phospholipid molecules in which proteins are embedded. These integral membrane proteins, which cross the bilayer once or several times, generally have a higher than average hydrophobicity and tend to aggregate. Detergents are needed to remove integral membrane proteins from the lipid bilayer and they have to be present during further chromatographic purification. Predominantly, four modes of HPLC have been used alone or in combination for the purification of integral membrane proteins. These are based on differences of proteins in size (size-exclusion chromatography, SEC), electrostatic interaction (ion-exchange chromatography, IEC), bioaffinity (bioaffinity chromatography, BAC) and hydrophobic interaction (reversed-phase chromatography, RPC, and hydrophobic-interaction chromatography, HIC). SEC, IEC, BAC and HIC are used under relatively mild conditions, and buffer systems generally contain a non-ionic detergent. RPC generally has a denaturing effect on the protein and should preferably be used for the purification of integral membrane proteins smaller than 50 kD.
生物膜的主要功能是在细胞和细胞器中对生物过程进行分隔。它们由磷脂分子双层组成，蛋白质嵌入其中。这些跨膜一次或多次的整合膜蛋白通常具有高于平均水平的疏水性，并且易于聚集。需要使用去污剂从脂质双层中去除整合膜蛋白，并且在进一步的色谱纯化过程中必须存在去污剂。主要有四种高效液相色谱模式单独或联合用于整合膜蛋白的纯化。这些模式基于蛋白质在大小（尺寸排阻色谱法，SEC）、静电相互作用（离子交换色谱法，IEC）、生物亲和力（生物亲和色谱法，BAC）和疏水相互作用（反相色谱法，RPC，以及疏水相互作用色谱法，HIC）方面的差异。SEC、IEC、BAC和HIC在相对温和的条件下使用，缓冲系统通常含有非离子去污剂。RPC通常对蛋白质具有变性作用，最好用于纯化小于50 kD的整合膜蛋白。
