STRUCTURAL BIOLOGY--EARLY EVENTS IN LIPOPROTEIN ASSEMBLY

富三酰基甘油脂蛋白（TAG- lp）的组装过程是一个复杂的过程，包括最初形成一个小的原始颗粒（120-200 AD），其核心是中性脂质（TAG和CE），随后加入TAG和磷脂（PL）使其大小增加到新生的VLDL （300-600 AD）。一旦合成载脂蛋白ob可遵循两条途径：如果有脂质，载脂蛋白ob在新生的富含TAG的颗粒上分泌；如果脂质不足，载脂蛋白ob被降解。乳腺源性C27细胞不产生载脂蛋白，不分泌脂质，也没有微粒体TAG转移蛋白（MTP）。这些细胞被用来研究仅由载脂蛋白ob初级序列指导的组装。当C127细胞转染c端截断apoB形式的cDNA时，它们在脂质缺乏状态下有效地分泌n端17%的apoB (B17)，但随着脂质逐渐增加，分泌apoB29、B32.5、B37和B41。我们发现B29结合PL和DAG， B32结合PL和TAG，而B32和B41之间的序列主要结合TAG。因此，PL和DAG结合的特异位点出现在B20 - B29之间的序列中，而特异的TAG位点出现在B32 - B41之间。B37和B41颗粒的结构分析表明载脂蛋白ob必须直接与核心相互作用。当油酸供应时，这些截断的形式更有效地分泌，如果脂质不足，则会降解。在组装过程中发现了几种中间折叠形式，这些形式结合了多种伴侣。一些伴侣蛋白似乎参与了早期折叠事件，而另一些则参与了针对非脂化、错误折叠形式的降解路径。对B41中潜在脂质结合序列的搜索表明，在B21和B41之间可能存在两亲性β链（AbetaS），可能以2至4股的形式排列。合成了一致的27aa两亲β片，并与碳氢/水界面紧密结合，使界面张力从50 Mn/M降至22 Mn/M。该薄片具有弹性，在压缩时不会从油/水界面移出。载脂蛋白ob结合TAG-LP疏水核心的理想特性。

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.