Structural Basis for Ceramide Recognition and Hydrolysis by Human Neutral Ceramidase
Structural Basis for Ceramide Recognition and Hydrolysis by Human Neutral Ceramidase
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DOI：
10.1016/j.str.2015.06.013
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发表时间：
2015-08-04
期刊：
影响因子：
5.7
通讯作者：
Hannun, Yusuf A.
中科院分区：
文献类型：
--
作者：
Airola, Michael V.;Allen, William J.;Hannun, Yusuf A.
中性神经酰胺酶（nCD酶）催化骨化相关脂质神经酰胺转化为鞘氨醇，鞘氨醇是增殖因子鞘氨醇-1-磷酸的前体。作为一种调节神经酰胺和鞘氨醇-1-磷酸平衡的酶，nCD酶正在成为癌症的治疗靶点。在这里，我们提出了2.6埃的晶体结构的人nCD酶与磷酸盐的复合物，揭示了一个惊人的，20埃深，疏水活性位点口袋稳定的真核生物特异性亚结构域不存在于细菌神经酰胺酶。利用灵活的配体对接，我们预测一个可能的结合模式神经酰胺，叠加密切与晶体学观察到的过渡态类似磷酸盐。我们的研究结果表明，nCDase使用一种新的催化策略，锌+依赖性酰胺酶，并产生神经酰胺特异性的空间排除鞘脂与庞大的头基和特异性识别的小羟基头基的神经酰胺。总之，这些数据为药物开发提供了基础，并为蛋白质如何识别生物活性脂质神经酰胺建立了共同的主题。
Neutral ceramidase (nCDase) catalyzes conversion of the apoptosis-associated lipid ceramide to sphingosine, the precursor for the proliferative factor sphingosine-1-phosphate. As an enzyme regulating the balance of ceramide and sphingosine-1-phosphate, nCDase is emerging as a therapeutic target for cancer. Here, we present the 2.6-angstrom crystal structure of human nCDase in complex with phosphate that reveals a striking, 20-angstrom deep, hydrophobic active site pocket stabilized by a eukaryotic-specific subdomain not present in bacterial ceramidases. Utilizing flexible ligand docking, we predict a likely binding mode for ceramide that superimposes closely with the crystallographically observed transition state analog phosphate. Our results suggest that nCDase uses a new catalytic strategy for Zn2+-dependent amidases, and generates ceramide specificity by sterically excluding sphingolipids with bulky headgroups and specifically recognizing the small hydroxyl head group of ceramide. Together, these data provide a foundation to aid drug development and establish common themes for how proteins recognize the bioactive lipid ceramide.