STRUCTURAL BIOLOGY--EARLY EVENTS IN LIPOPROTEIN ASSEMBLY

富含三酰甘油的脂蛋白(TAG-LP)的组装过程是一个复杂的过程，首先形成一个具有中性脂(TAG和CE)核心的原始小颗粒(120-200AD)，然后加入TAG和磷脂(PL)，使其大小增加到新生的极低密度脂蛋白(300-600AD)的尺寸。一旦合成载脂蛋白B可以遵循两条途径：如果有脂类，载脂蛋白B被分泌在新生的富含标签的颗粒上-如果脂类缺乏，载脂蛋白B被降解。乳腺来源的C27细胞不产生载脂蛋白，不分泌脂质，也不表达微粒体标签转移蛋白(MTP)。这些细胞被用来研究只由apoB的一级序列指导的组装。将C端截短的apoB基因导入C127细胞后，C127细胞能有效地分泌N端17%的apoB(B17)，而apoB29、B32.5、B37和B41随着脂类含量的递增而分泌。我们发现B29结合了PL和DAG，B32结合了PL和Tag，而B32和B41之间的序列主要结合了Tag。因此，PL和DAG结合的特定位点出现在B20和B29之间的序列中，而特定标签位点出现在B32到B41之间。对B37和B41粒子的结构分析表明，apoB必须与核心直接作用。当供应油酸盐时，这些截短形式的脂肪分泌更有效，如果脂质不足，这些截短形式的脂肪就会被降解。在组装过程中已经确定了几种中间折叠形式，这些形式结合了各种伴侣。一些伴侣似乎参与了早期的折叠事件，另一些则针对未脂化的、错误折叠的形式朝向降解路径/对B41中潜在的脂结合序列的搜索表明，在B21和B41之间存在两亲性β链(AbetaS)，可能以2至4条链的形式组织。合成了一个公认的27个氨基酸的两亲性β链，并将其强烈地结合到烃/水界面上，使界面张力从50 mN/M降至22 mN/M。该链具有弹性，压缩时不能从油/水界面移开。ApoB与Tag-LP疏水核心结合的理想性质。

The process of assembly of triacylglycerol rich lipoproteins (TAG-LP) is a complicated process involving the initial formation of a small primordial (120-200 AD) particle with a neutral lipid (TAG and CE) core and a later process which adds TAG and phospholipid (PL) to increase the size to that of a nascent VLDL (300-600 A D). Once synthesized apoB can follow 2 pathways: if lipid is available, apoB is secreted on nascent TAG rich particles-if lipid is deficient, apoB is degraded. Mammary derived C27 cells make no apolipoproteins, secreted no lipid and have no microsomal TAG transfer protein (MTP). These cells are used to study assembly directed only by the primary sequence of apoB. When C127 cells are transfected with cDNA for C-terminal truncated apoB forms, they efficiently secrete the N-terminal 17% of apoB (B17) in a lipid poor state but secrete apoB29, B32.5, B37, and B41 with progressively increasing amounts of lipids. We show that B29 binds PL and DAG, B32 binds PL and TAG, while the sequences between B32 and B41 bind mainly TAG. Thus, specific sites for PL and DAG binding appear in the sequence between B20 and B29 while specific TAG sites occur from B32 to B41. Structural analysis of B37 and B41 particles indicated that apoB must interact directly with the core. These truncated forms are secreted more efficiently when oleate is supplied and degraded if lipids are deficient. Several intermediate folding forms have been identified in the assembly process and these forms bind a variety of chaperones. Some chaperones appear to be involved in early folding events and others in targeting unlipidated, misfolded forms toward the degradation path/ A search for potential lipid binding sequences in B41 indicates that there are amphipathic beta strands (AbetaS) located between B21 and B41 probably organized in sheets of 2 to 4 strands. A consensus 27 aa amphipathic beta sheet was synthesized and bound avidly to a hydrocarbon/water interface lowering the interfacial tension from 50 to 22 Mn/M. The sheet bound elastically and could not be displaced from the oil/water interface when compressed. An ideal property for apoB binding to the hydrophobic core of TAG-LP.