Forte T M, Bielicki J K, Knoff L, McCall M R
Department of Molecular and Nuclear Medicine, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.
J Lipid Res. 1996 May;37(5):1076-85.
Apolipoprotein A-I (apoA-I) incubated with CHO cells assembles three major nascent lipid complexes with diameters of 7.3, 9, and 11 nm. Previous studies suggested that the smaller nascent particles were precursors for the larger nascent ones. To test this hypothesis, the 7.3, 9, and 11 nm apoA-I-lipid complexes formed by incubating CHO cells with lipid-free apoA-I were isolated and subsequently each subpopulation was re-incubated with cells in the absence of other subpopulations. The physical-chemical characteristics of each subpopulation were examined before and after re-incubation in an effort to understand relationships. if any, between the different nascent complexes. The 7.3, 9, and 11 nm complexes were unique in that each of the particles had pre-alpha mobility on agarose gels: this rapid migration was not altered by re-incubation with cells. Protein crosslinking studies indicated that the 7.3, 9, and 11 nm complexes possessed 2, 3, and 4 apoA-I molecules per complex, respectively; it is unlikely that the size of the particle and number of apoA-I molecules per particle played a role in the increased negative charge of the particles. The present study shows that smaller particles did not give rise to larger ones upon re-incubation with cells. Rather, the 11 and 9 nm particles both generated smaller discs (the 11 nm giving rise primarily to 9 nm discs and the 9 nm complex giving rise to 7.3 nm discs) suggesting that, during incubation with cells, the complexes are destabilized and remodeled into smaller, not larger, complexes. Surprisingly, the 7.3 nm complexes during re-incubation with cells were extremely stable and did not undergo size alteration. When the 7.3 nm particles were incubated with additional small quantities of lipid-free apoA-I (1-2 microgram/ml), larger discoidal complexes were generated suggesting that the formation of larger particles may be driven by the availability of lipid-free apoA-I.
与中国仓鼠卵巢细胞（CHO细胞）一起孵育的载脂蛋白A-I（apoA-I）会组装出三种主要的新生脂质复合物，其直径分别为7.3、9和11纳米。先前的研究表明，较小的新生颗粒是较大新生颗粒的前体。为了验证这一假设，将通过用无脂质的apoA-I孵育CHO细胞形成的7.3、9和11纳米的apoA-I-脂质复合物分离出来，随后在没有其他亚群的情况下，将每个亚群分别与细胞重新孵育。在重新孵育前后检查每个亚群的物理化学特性，以试图了解不同新生复合物之间的关系（如果有的话）。7.3、9和11纳米的复合物具有独特之处，即每个颗粒在琼脂糖凝胶上都具有前α迁移率；这种快速迁移不会因与细胞重新孵育而改变。蛋白质交联研究表明，7.3、9和11纳米的复合物每复合物分别含有2、3和4个apoA-I分子；颗粒大小和每个颗粒中apoA-I分子的数量不太可能在颗粒负电荷增加中起作用。本研究表明，较小的颗粒在与细胞重新孵育后不会产生较大的颗粒。相反，11纳米和9纳米的颗粒都会产生较小的盘状物（11纳米的主要产生9纳米的盘状物，9纳米的复合物产生7.3纳米的盘状物），这表明在与细胞孵育期间，复合物不稳定并重塑为更小而非更大的复合物。令人惊讶的是，7.3纳米的复合物在与细胞重新孵育期间极其稳定，且大小没有改变。当7.3纳米的颗粒与少量额外的无脂质apoA-I（1-2微克/毫升）一起孵育时，会产生更大的盘状复合物，这表明更大颗粒的形成可能由无脂质apoA-I的可用性驱动。
