Evidence for triacylglycerol synthesis in the lumen of microsomes via a lipolysis-esterification pathway involving carnitine acyltransferases
Evidence for triacylglycerol synthesis in the lumen of microsomes via a lipolysis-esterification pathway involving carnitine acyltransferases
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DOI：
10.1074/jbc.274.50.35577
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发表时间：
1999-12-10
影响因子：
4.8
通讯作者：
Clarke, D
中科院分区：
文献类型：
--
作者：
Abo-Hashema, KAH;Cake, MH;Clarke, D
在这项研究中，已经鉴定了内质网腔内合成三酰甘油（TAG）的途径，使用已经通过耗尽其内源性底物进行预处理的微粒体，然后将它们与含有CoASH和Mg 2+的生物素化磷脂酰丝氨酸脂质体融合。将这些融合的微粒体与三[H-3]油酰甘油和[C-14]油酰-CoA孵育产生微粒体相关的三酰甘油，其耐大量洗涤且具有接近2：1的[3 H]：[14 C]比。数据表明前体三[H-3]油酰甘油被微粒体脂肪酶水解为膜结合的二[3 H]油酰甘油，随后用鲁米那[C-14]油酰-CoA再酯化。TAG在微粒体内的蓄积，即使在显性二酰基甘油酰基转移酶（DGAT I）无活性时，也与微粒体腔内存在潜伏性二酰基甘油酰基转移酶（DGAT II）一致。此外，由于TAG的管腔合成是肉毒碱依赖性的，并被格列本脲（一种有效的肉毒碱酰基转移酶抑制剂）显著降低，因此微粒体肉毒碱酰基转移酶似乎对于将[C-14]油酰CoA运输到微粒体管腔中以随后掺入新合成的TAG是必不可少的。因此，这项研究提供了第一个直接证明的酶促过程，导致鲁米那三酰甘油的合成，这是一个非常低密度脂蛋白的主要组成部分。
In this study a pathway for the synthesis of triacylglycerol (TAG) within the lumen of the endoplasmic reticulum has been identified, using microsomes that had been preconditioned by depleting their endogenous substrates and then fusing them with biotinylated phosphatidylserine liposomes containing CoASH and Mg2+ Incubating these fused microsomes with tri[H-3] oleoylglycerol and [C-14]oleoyl-CoA yielded microsome-associated triacylglycerol, which resisted extensive washing and had a [3H]:[14C] ratio close to 2:1, The data suggest that the precursor tri[H-3]oleoylglycerol was hydrolyzed by microsomal lipase to membrane-bound di[3H]oleoylglycerol and subsequently re-esterified with luminal [C-14]oleoyl-CoA. The accumulation of TAG within the microsomes, even when overt diacylglycerol acyltransferase (DGAT I) was inactive, is consistent with the existence of a latent diacylglycerol acyltransferase (DGAT II) within the microsomal lumen. Moreover, because luminal synthesis of TAG was carnitine-dependent and markedly reduced by glybenclamide, a potent carnitine acyltransferase inhibitor, microsomal carnitine acyltransferase appears to be essential for trafficking the [C-14]oleoyl CoA into the microsomal lumen for subsequent incorporation into newly synthesized TAG. This study thus provides the first direct demonstration of an enzymatic process leading to the synthesis of luminal triacylglycerol, which is a major component of very low density lipoproteins.