Structural Basis of Ubiquitin Recognition by a Bacterial Ovarian Tumor Deubiquitinase LotA
Structural Basis of Ubiquitin Recognition by a Bacterial Ovarian Tumor Deubiquitinase LotA
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DOI：
10.1128/jb.00376-21
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发表时间：
2021-10
影响因子：
3.2
通讯作者：
N. Takekawa;T. Kubori;Tomoya Iwai;H. Nagai;K. Imada
中科院分区：
文献类型：
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作者：
N. Takekawa;T. Kubori;Tomoya Iwai;H. Nagai;K. Imada
为了成功地定殖，一些病原菌劫持宿主泛素系统。军团菌OTU-like-DUBs（Lot-DUBs）是在军团菌的效应蛋白中发现的新型细菌去泛素化酶。嗜肺菌LotA是Lot-DUB的成员，具有两个OTU域（OTU 1和OTU 2）。摘要病原菌通过与宿主细胞的密切相互作用获得了大量的真核生物蛋白质样蛋白。作为泛素连接酶和去泛素化酶（DUB）起作用的细菌效应蛋白是这种分子模拟的显著实例。LotA是嗜肺军团菌的效应子，属于卵巢肿瘤（OTU）超家族，通过DUB活性调节多种泛素信号。LotA含有两个具有不同反应性的OTU结构域;第一个负责K6连接的泛素链的切割，第二个显示出对长链泛素的不常见的偏好。在这里，我们报告了LotA（LotAM）的中间结构域的晶体结构，其中包含第二个OTU结构域。LotAM由两个不同的亚结构域组成，一个是与人OTU DUB具有高度结构相似性的催化结构域，另一个是仅在军团菌OTU DUB中特征性保守的延伸螺旋叶（EHL）结构域。LotAM与ubiquitin的对接模拟表明，LotAM的EHL和ubiquitin的C-末端区域之间的疏水和静电相互作用是ubiquitin与LotAM结合的关键。基于结构的诱变表明，在被称为“螺旋臂”的特征性短螺旋片段中的酸性残基对于LotAM的酶活性是必需的。三种军团菌OTU DUB（LotA、LotB和LotC）的EHL结构域共享“螺旋臂”结构，表明EHL结构域将Lot-OTU定义为独特类别的DUB。重要性为了成功地定殖，一些病原菌劫持宿主泛素系统。军团菌OTU-like-DUBs（Lot-DUBs）是在军团菌的效应蛋白中发现的新型细菌去泛素化酶。嗜肺菌LotA是Lot-DUB的成员，具有两个OTU域（OTU 1和OTU 2）。我们确定了LotA（LotAM）的中间片段的结构，其中包括OTU 2。LotAM由保守的催化结构域和军团菌OTUs特异性EHL结构域组成。与泛素的对接模拟和突变分析表明，在EHL的酸性表面是必不可少的酶活性。EHL的结构不同于其他批次DUB，表明EHL的变化与每个DUB的可变切割特异性有关。
To successfully colonize, some pathogenic bacteria hijack the host ubiquitin system. Legionella OTU-like-DUBs (Lot-DUBs) are novel bacterial deubiquitinases found in effector proteins of L. pneumophila. LotA is a member of Lot-DUBs and has two OTU domains (OTU1 and OTU2). ABSTRACT Pathogenic bacteria have acquired a vast array of eukaryotic-protein-like proteins via intimate interaction with host cells. Bacterial effector proteins that function as ubiquitin ligases and deubiquitinases (DUBs) are remarkable examples of such molecular mimicry. LotA, a Legionella pneumophila effector, belongs to the ovarian tumor (OTU) superfamily, which regulates diverse ubiquitin signals by their DUB activities. LotA harbors two OTU domains that have distinct reactivities; the first one is responsible for the cleavage of the K6-linked ubiquitin chain, and the second one shows an uncommon preference for long chains of ubiquitin. Here, we report the crystal structure of a middle domain of LotA (LotAM), which contains the second OTU domain. LotAM consists of two distinct subdomains, a catalytic domain having high structural similarity with human OTU DUBs and an extended helical lobe (EHL) domain, which is characteristically conserved only in Legionella OTU DUBs. The docking simulation of LotAM with ubiquitin suggested that hydrophobic and electrostatic interactions between the EHL of LotAM and the C-terminal region of ubiquitin are crucial for the binding of ubiquitin to LotAM. The structure-based mutagenesis demonstrated that the acidic residue in the characteristic short helical segment termed the “helical arm” is essential for the enzymatic activity of LotAM. The EHL domain of the three Legionella OTU DUBs, LotA, LotB, and LotC, share the “helical arm” structure, suggesting that the EHL domain defines the Lot-OTUs as a unique class of DUBs. IMPORTANCE To successfully colonize, some pathogenic bacteria hijack the host ubiquitin system. Legionella OTU-like-DUBs (Lot-DUBs) are novel bacterial deubiquitinases found in effector proteins of L. pneumophila. LotA is a member of Lot-DUBs and has two OTU domains (OTU1 and OTU2). We determined the structure of a middle fragment of LotA (LotAM), which includes OTU2. LotAM consists of the conserved catalytic domain and the Legionella OTUs-specific EHL domain. The docking simulation with ubiquitin and the mutational analysis suggested that the acidic surface in the EHL is essential for enzymatic activity. The structure of the EHL differs from those of other Lot-DUBs, suggesting that the variation of the EHL is related to the variable cleaving specificity of each DUB.