Cryo-EM structures of human fucosidase FucA1 reveal insight into substrate recognition and catalysis.
Cryo-EM structures of human fucosidase FucA1 reveal insight into substrate recognition and catalysis.
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DOI：
10.1016/j.str.2022.07.001
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发表时间：
2022-10-06
期刊：
影响因子：
5.7
通讯作者：
Davies, Gideon J.
中科院分区：
文献类型：
--
作者：
Armstrong, Zachary;Meek, Richard W.;Wu, Liang;Blaza, James N.;Davies, Gideon J.
从岩藻糖基化糖缀合物酶促水解α-L-岩藻糖在细菌感染和神经退行性溶酶体贮积症岩藻糖苷沉积症中是必然的。由于缺乏任何动物岩藻糖苷酶的三维结构数据，在药物设计方面，对人类α-L-岩藻糖苷酶催化作用的理解受到阻碍。在这里，我们使用冷冻电子显微镜（cryo-EM）来确定人溶酶体α-L-岩藻糖苷酶（FucA 1）的结构，无论是在未配体状态还是与抑制剂脱氧岩野尻霉素的复合物中。这些结构，在2.49 nm分辨率下确定，揭示了FucA 1的同源四聚体结构，催化中心的结构，以及自然群体变异和致病突变的位置。此外，这项工作最终确定了迄今为止有争议的催化酸/碱的身份为天冬氨酸-276，代表了经典谷氨酸/碱残基和之前提出的谷氨酸残基的转变。这些发现进一步加深了我们对FucA 1如何在健康和疾病中发挥作用的理解。FucA 1的Cryo-EM结构解决了关于催化机制的争论天冬氨酸276，这是所有动物同系物的保守，是催化酸/碱一种错误折叠的FucA 1疾病变体（S150 F）通过亚氨基糖抑制剂稳定人溶酶体岩藻糖苷酶（FucA 1）分解岩藻糖基化聚糖。酶的异常导致岩藻糖苷沉积症。Armstrong等人使用单颗粒冷冻电镜来解决这种酶的争议机制。他们还证明了脱氧岩野尻霉素的错误折叠变体的稳定性，并确定了FucA 1如何容纳这种抑制剂。
Enzymatic hydrolysis of α-L-fucose from fucosylated glycoconjugates is consequential in bacterial infections and the neurodegenerative lysosomal storage disorder fucosidosis. Understanding human α-L-fucosidase catalysis, in an effort toward drug design, has been hindered by the absence of three-dimensional structural data for any animal fucosidase. Here, we have used cryoelectron microscopy (cryo-EM) to determine the structure of human lysosomal α-L-fucosidase (FucA1) in both an unliganded state and in complex with the inhibitor deoxyfuconojirimycin. These structures, determined at 2.49 Å resolution, reveal the homotetrameric structure of FucA1, the architecture of the catalytic center, and the location of both natural population variations and disease-causing mutations. Furthermore, this work has conclusively identified the hitherto contentious identity of the catalytic acid/base as aspartate-276, representing a shift from both the canonical glutamate acid/base residue and a previously proposed glutamate residue. These findings have furthered our understanding of how FucA1 functions in both health and disease. Cryo-EM structure of FucA1 resolves debate over the catalytic mechanism Aspartate 276, which is conserved for all animal homologs, is the catalytic acid/base A misfolded FucA1 disease variant (S150F) is stabilized by an iminosugar inhibitor Human lysosomal fucosidase (FucA1) recycles fucosylated glycans. Aberrations in the enzyme result in fucosidosis. Armstrong et al. use single-particle cryo-EM to resolve the contentious mechanism of this enzyme. They also demonstrate stabilization of a misfolded variant by deoxyfuconojirimycin and determine how FucA1 accommodates this inhibitor.
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