Carboxylesterase 3 (EC 3.1.1.1) is a major adipocyte lipase
Carboxylesterase 3 (EC 3.1.1.1) is a major adipocyte lipase
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DOI：
10.1074/jbc.m400541200
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发表时间：
2004-09-24
影响因子：
4.8
通讯作者：
Mitchell, GA
中科院分区：
文献类型：
--
作者：
Soni, KG;Lehner, R;Mitchell, GA
甘油三酯的水解是白色脂肪组织（WAT）能量稳态的核心。以前认为激素敏感脂肪酶（HSL）介导WAT中的所有脂解。令人惊讶的是，HSL缺陷小鼠显示出活跃的HSL非依赖性脂解，表明其他脂肪酶也介导甘油三酯水解。为了阐明这一点，我们使用功能蛋白质组学来检测小鼠WAT中的非HSL脂肪酶。在腹腔内WAT的细胞分级分离后，大多数非HSL中性脂肪酶活性位于100，000 x g的下层和脂肪饼级分中。通过油酸连接的琼脂糖层析的infranatant，然后在3-[（3-cholamidopropyl）dimethylammonio]-1-propanesulfonic acid梯度洗脱，我们确定了两个峰的酯酶活性使用对硝基苯基丁酸酯作为底物。其中一个峰包含大部分脂肪酶活性。在相应的级分中，凝胶渗透色谱法和SDS-PAGE，然后对切下的考马斯蓝染色的肽进行串联质谱分析，揭示了羧酸酯酶3（三酰甘油水解酶（TGH）; EC 3.1.1.1）。TGH也是WAT脂肪饼提取物的主要脂肪酶。部分纯化的WAT TGH具有脂肪酶活性以及较小但可检测到的中性胆固醇酯水解酶活性。WAT的亚细胞组分的Western印迹和体外脂肪细胞分化后成纤维细胞的共聚焦显微镜观察结果与TGH在内质网、胞质溶胶和脂滴中的分布一致。TGH在体外WAT中负责非HSL脂肪酶活性的主要部分，并且可以介导脂肪细胞中的部分或全部不依赖于HSL的脂解。
Hydrolysis of triglycerides is central to energy homeostasis in white adipose tissue (WAT). Hormone-sensitive lipase (HSL) was previously felt to mediate all lipolysis in WAT. Surprisingly, HSL-deficient mice show active HSL-independent lipolysis, suggesting that other lipase(s) also mediate triglyceride hydrolysis. To clarify this, we used functional proteomics to detect non-HSL lipase(s) in mouse WAT. After cell fractionation of intra-abdominal WAT, most non-HSL neutral lipase activity is localized in the 100,000 x g infranatant and fat cake fractions. By oleic acid-linked agarose chromatography of infranatant followed by elution in a 3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonic acid gradient, we identified two peaks of esterase activity using p-nitrophenyl butyrate as a substrate. One of the peaks contained most of the lipase activity. In the corresponding fractions, gel permeation chromatography and SDS-PAGE, followed by tandem mass spectrometric analysis of excised Coomassie Blue-stained peptides, revealed carboxylesterase 3 (triacylglycerol hydrolase (TGH); EC 3.1.1.1). TGH is also the principle lipase of WAT fat cake extracts. Partially purified WAT TGH had lipase activity as well as lesser but detectable neutral cholesteryl ester hydrolase activity. Western blotting of subcellular fractions of WAT and confocal microscopy of fibroblasts following in vitro adipocytic differentiation are consistent with a distribution of TGH to endoplasmic reticulum, cytosol, and the lipid droplet. TGH is responsible for a major part of non-HSL lipase activity in WAT in vitro and may mediate some or all HSL-independent lipolysis in adipocytes.