Structure of human pancreatic lipase-related protein 2 with the lid in an open conformation
Structure of human pancreatic lipase-related protein 2 with the lid in an open conformation
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DOI：
10.1021/bi8005576
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发表时间：
2008-09-09
期刊：
影响因子：
2.9
通讯作者：
Carriere, Frederic
中科院分区：
文献类型：
--
作者：
Eydoux, Cecilia;Spinelli, Silvia;Carriere, Frederic
进入胰腺脂肪酶(PL)活性部位的途径是由一个表面环，即盖子控制的，通常只有在添加脂类或两亲分子时才会发生构象变化。具有开放和功能构象的PL的结构需要与两亲分子共结晶。在这里，我们报道了野生型和非糖基化的人胰腺脂肪酶相关蛋白2(HPLRP2)的两种晶体结构，在没有两亲分子的情况下，盖子呈开放构象。这些独立求解的结构惊人地相似，在电子密度图中，一些盖子的残基定义不佳。然而，该盖子的开放构象与以前在经典配体PL中观察到的不同，表明HPLRP2具有不同的动力学性质。在这里，我们表明，HPLRP2直接被E600抑制，不存在界面激活，并优先作用于形成单体或分散在溶液中的小聚集体(胶束)的底物，如单甘酯、磷脂和半乳糖脂，而经典PL显示出相反的特性和对形成水-油界面的不溶性底物，如甘油三酯和双甘酯的高度专一性。这些生化特性表明，HPLPP2的盖子很可能在溶液中自发采用晶体结构中观察到的开放构象。这种开放的构象产生了一个大的空腔，能够容纳半乳糖脂的双半乳糖极头，类似于以前在豚鼠PLRP2活性部位观察到的空腔，但没有经典的PL。HPLRP2的大部分结构和动力学性质与大鼠PLRP2的不同，大鼠PLRP2的结构是通过封闭构象的盖子获得的。我们的发现说明了LID在决定脂肪酶底物专一性和作用机制方面的重要作用。
Access to the active site of pancreatic lipase (PL) is controlled by a surface loop, the lid, which normally undergoes conformational changes only upon addition of lipids or amphiphiles. Structures of PL with their lids in the open and functional conformation have required cocrystallization with amphiphiles. Here we report two crystal structures of wild-type and unglycosylated human pancreatic lipase-related protein 2 (HPLRP2) with the lid in an open conformation in the absence of amphiphiles. These structures solved independently are strikingly similar, with some residues of the lid being poorly defined in the electron-density map. The open conformation of the lid is however different from that previously observed in classical liganded PL, suggesting different kinetic properties for HPLRP2. Here we show that the HPLRP2 is directly inhibited by E600, does not present interfacial activation, and acts preferentially on substrates forming monomers or small aggregates (micelles) dispersed in solution like monoglycerides, phospholipids and galactolipids, whereas classical PL displays reverse properties and a high specificity for unsoluble substrates like triglycerides and diglycerides forming oil-in-water interfaces. These biochemical properties imply that the lid of HPLPP2 is likely to spontaneously adopt in solution the open conformation observed in the crystal structure. This open conformation generates a large cavity capable of accommodating the digalactose polar head of galactolipids, similar to that previously observed in the active site of the guinea pig PLRP2, but absent from the classical PL. Most of the structural and kinetic properties of HPLRP2 were found to be different from those of rat PLRP2, the structure of which was previously obtained with the lid in a closed conformation. Our findings illustrate the essential role of the lid in determining the substrate specificity and the mechanism of action of lipases.