Khatib Abdallah, Arhab Yani, Bentebibel Assia, Abousalham Abdelkarim, Noiriel Alexandre
Institut de Chimie et de Biochimie Moléculaires et Supramoléculaires UMR 5246 CNRS, Organisation et Dynamique des Membranes Biologiques, Université Lyon 1, Villeurbanne, France.
PLoS One. 2016 Jan 8;11(1):e0145806. doi: 10.1371/journal.pone.0145806. eCollection 2016.
Comparative Gene Identification-58 (CGI-58) is a widespread protein found in animals and plants. This protein has been shown to participate in lipolysis in mice and humans by activating Adipose triglyceride lipase (ATGL), the initial enzyme responsible for the triacylglycerol (TAG) catabolism cascade. Human mutation of CGI-58 is the cause of Chanarin-Dorfman syndrome, an orphan disease characterized by a systemic accumulation of TAG which engenders tissue disorders. The CGI-58 protein has also been shown to participate in neutral lipid metabolism in plants and, in this case, a mutation again provokes TAG accumulation. Although its roles as an ATGL coactivator and in lipid metabolism are quite clear, the catalytic activity of CGI-58 is still in question. The acyltransferase activities of CGI-58 have been speculated about, reported or even dismissed and experimental evidence that CGI-58 expressed in E. coli possesses an unambiguous catalytic activity is still lacking. To address this problem, we developed a new set of plasmids and site-directed mutants to elucidate the in vivo effects of CGI-58 expression on lipid metabolism in E. coli. By analyzing the lipid composition in selected E. coli strains expressing CGI-58 proteins, and by reinvestigating enzymatic tests with adequate controls, we show here that recombinant plant CGI-58 has none of the proposed activities previously described. Recombinant plant and mouse CGI-58 both lack acyltransferase activity towards either lysophosphatidylglycerol or lysophosphatidic acid to form phosphatidylglycerol or phosphatidic acid and recombinant plant CGI-58 does not catalyze TAG or phospholipid hydrolysis. However, expression of recombinant plant CGI-58, but not mouse CGI-58, led to a decrease in phosphatidylglycerol in all strains of E. coli tested, and a mutation of the putative catalytic residues restored a wild-type phenotype. The potential activities of plant CGI-58 are subsequently discussed.
比较基因识别蛋白58（CGI-58）是一种在动植物中广泛存在的蛋白质。该蛋白已被证明通过激活脂肪甘油三酯脂肪酶（ATGL）参与小鼠和人类的脂肪分解，ATGL是负责三酰甘油（TAG）分解代谢级联反应的起始酶。人类CGI-58突变是钱纳林-多夫曼综合征的病因，这是一种罕见病，其特征是TAG在全身积累，导致组织功能紊乱。CGI-58蛋白也被证明参与植物的中性脂质代谢，在这种情况下，突变同样会导致TAG积累。尽管其作为ATGL共激活因子的作用以及在脂质代谢中的作用相当明确，但CGI-58的催化活性仍存在疑问。关于CGI-58的酰基转移酶活性已有推测、报道甚至否定，但仍缺乏在大肠杆菌中表达的CGI-58具有明确催化活性的实验证据。为了解决这个问题，我们开发了一套新的质粒和定点突变体，以阐明CGI-58表达对大肠杆菌脂质代谢的体内影响。通过分析表达CGI-58蛋白的选定大肠杆菌菌株中的脂质组成，并通过重新进行有适当对照的酶促试验，我们在此表明重组植物CGI-58不具有先前描述的任何假定活性。重组植物和小鼠CGI-58都缺乏对溶血磷脂酰甘油或溶血磷脂酸形成磷脂酰甘油或磷脂酸的酰基转移酶活性，并且重组植物CGI-58不催化TAG或磷脂水解。然而，重组植物CGI-58（而非小鼠CGI-58）的表达导致所有测试的大肠杆菌菌株中磷脂酰甘油减少，并且假定催化残基的突变恢复了野生型表型。随后讨论了植物CGI-58的潜在活性。
