Roussel A, Yang Y, Ferrato F, Verger R, Cambillau C, Lowe M
Architecture et Fonction des Macromolécules Biologiques, CNRS-IFR1 UPR 9039, 31 Chemin Joseph Aiguier, 13402 Marseille cedex 20, France.
J Biol Chem. 1998 Nov 27;273(48):32121-8. doi: 10.1074/jbc.273.48.32121.
The pancreas expresses several members of the lipase gene family including pancreatic triglyceride lipase (PTL) and two homologous proteins, pancreatic lipase-related proteins 1 and 2 (PLRP1 and PLRP2). Despite their similar amino acid sequences, PTL, PLRP1, and PLRP2 differ in important kinetic properties. PLRP1 has no known activity. PTL and PLRP2 differ in substrate specificity, bile acid inhibition, colipase requirement, and interfacial activation. To begin understanding the structural explanations for these functional differences, we solved the crystal structure of rat (r)PLRP2 and further characterized its kinetic properties. The 1.8 A structure of rPLRP2, like the tertiary structure of human PTL, has a globular N-terminal domain and a beta-sandwich C-terminal domain. The lid domain occupied the closed position, suggesting that rPLRP2 should show interfacial activation. When we reexamined this issue with tripropionin as substrate, rPLRP2 exhibited interfacial activation. Because the active site topology of rPLRP2 resembled that of human PTL, we predicted and demonstrated that the lipase inhibitors E600 and tetrahydrolipstatin inhibit rPLRP2. Although PTL and rPLRP2 have similar active sites, rPLRP2 has a broader substrate specificity that we confirmed using a monolayer technique. With this assay, we showed for the first time that rPLRP2 prefers phosphatidylglycerol and ethanolamine over phosphatidylcholine. In summary, we confirmed and extended the observation that PLRP2 lipases have a broader substrate specificity than PTL, we demonstrated that PLRP2 lipases show interfacial activation, and we solved the first crystal structure of a PLRP2 lipase that contains a lid domain.
胰腺表达脂肪酶基因家族的多个成员，包括胰腺甘油三酯脂肪酶（PTL）以及两种同源蛋白，即胰腺脂肪酶相关蛋白1和2（PLRP1和PLRP2）。尽管它们的氨基酸序列相似，但PTL、PLRP1和PLRP2在重要的动力学特性方面存在差异。PLRP1尚无已知活性。PTL和PLRP2在底物特异性、胆汁酸抑制作用、辅脂酶需求以及界面激活方面有所不同。为了开始理解这些功能差异的结构解释，我们解析了大鼠（r）PLRP2的晶体结构，并进一步表征了其动力学特性。rPLRP2的1.8埃结构与人类PTL的三级结构一样，具有一个球状的N端结构域和一个β-折叠三明治C端结构域。盖子结构域处于关闭位置，这表明rPLRP2应表现出界面激活。当我们以三丙酸甘油酯为底物重新审视这个问题时，rPLRP2表现出界面激活。由于rPLRP2的活性位点拓扑结构与人类PTL相似，我们预测并证明脂肪酶抑制剂E600和四氢脂抑素可抑制rPLRP2。尽管PTL和rPLRP2具有相似的活性位点，但rPLRP2具有更广泛的底物特异性，我们使用单层技术证实了这一点。通过该测定法，我们首次表明rPLRP2对磷脂酰甘油和乙醇胺的偏好超过磷脂酰胆碱。总之，我们证实并扩展了以下观察结果：PLRP2脂肪酶比PTL具有更广泛的底物特异性，我们证明PLRP2脂肪酶表现出界面激活，并且我们解析了首个包含盖子结构域的PLRP2脂肪酶的晶体结构。
