CryoEM structure at 9 Å resolution of an adenovirus vector targeted to hematopoietic cells
CryoEM structure at 9 Å resolution of an adenovirus vector targeted to hematopoietic cells
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DOI：
10.1016/j.jmb.2005.04.034
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发表时间：
2005-06-10
影响因子：
5.6
通讯作者：
Stewart, PL
中科院分区：
文献类型：
--
作者：
Saban, SD;Nepomuceno, RR;Stewart, PL
我们报告了一个亚纳米分辨率冷冻电子显微镜（cryoEM）结构分析的腺病毒载体，Ad 35 F，由一个腺病毒5型（Ad 5）衣壳假型与Ad 35纤维。该载体通过其纤维蛋白与补体调节蛋白家族成员CD 46的结合转导人造血细胞。数据采集和图像处理方面的重大进步使得分辨率与早期结构相比有了显着提高。通过对接六邻体和五邻体基础衣壳蛋白的晶体结构来增强cryoEM密度的分析。观察到晶体结构中缺失的六邻体残基的CryoEM密度，所述六邻体残基包括高变区和中和单克隆抗体的表位。在五邻体基底内，观察到晶体结构中缺失的整合素结合RGD环和五邻体基底一侧可变环的柔性P带的密度。Ad 35纤维是柔性的，与第三个P-螺旋重复序列中的序列插入一致。在内衣壳上，在六邻体的基部和五邻体基部下方显示表面密度。修正后的模型中的病毒体蛋白IX。定义明确的密度被分配到一个保守的结构域在N末端的蛋白IX所需的纳入病毒体。对于蛋白IX的C-末端结构域，提出了两种替代构象，要么结合在衣壳表面上，要么远离衣壳延伸。该模型与C末端对插入配体的耐受性及其在载体重靶向中的潜在用途一致。这种结构的研究增加了我们的知识，广告衣壳装配，抗体中和机制，并可能有助于进一步改善基因传递到重要的人类细胞类型。(c)2005爱思唯尔有限公司保留所有权利。
We report a sub-nanometer resolution cryo-electron microscopy (cryoEM) structural analysis of an adenoviral vector, Ad35F, comprised of an adenovirus type 5 (Ad5) capsid pseudo-typed with an Ad35 fiber. This vector transduces human hematopoietic cells via association of its fiber protein with CD46, a member of the complement regulatory protein family. Major advances in data acquisition and image, processing allowed a significant improvement in resolution compared to earlier structures. Analysis of the cryoEM density was enhanced by docking the crystal structures of both the hexon and penton base capsid proteins. CryoEM density was observed for hexon residues missing from the crystal structure that include hypervariable regions and the epitope of a neutralizing monoclonal antibody. Within the penton base, density was observed for the integrin-binding RGD loop missing from the crystal structure and for the flexible P ribbon of the variable loop on the side of the penton base. The Ad35 fiber is flexible, consistent with the sequence insert in the third P-spiral repeat. On the inner capsid surface density is revealed at the base of the hexons and below the penton base. A revised model is presented for protein IX within the virion. Well-defined density was assigned to a conserved domain in the N terminus of protein IX required for incorporation into the virion. For the C-terminal domain of protein IX two alternate conformations are proposed, either binding on the capsid surface or extending away from the capsid. This model is consistent with the tolerance of the C terminus for inserted ligands and its potential use in vector retargeting. This structural study increases our knowledge of Ad capsid assembly, antibody neutralization mechanisms, and may aid further improvements in gene delivery to important human cell types. (c) 2005 Elsevier Ltd. All rights reserved.