Xie Qing, Spear John M, Noble Alex J, Sousa Duncan R, Meyer Nancy L, Davulcu Omar, Zhang Fuming, Linhardt Robert J, Stagg Scott M, Chapman Michael S
Department of Biochemistry & Molecular Biology, School of Medicine, Oregon Health & Science University, Portland, OR 97239-3098, USA.
Institute of Molecular Biophysics, Florida State University, 91 Chieftan Way, Tallahassee, FL 32306-4380, USA.
Mol Ther Methods Clin Dev. 2017 Mar 8;5:1-12. doi: 10.1016/j.omtm.2017.02.004. eCollection 2017 Jun 16.
Atomic structures of adeno-associated virus (AAV)-DJ, alone and in complex with fondaparinux, have been determined by cryoelectron microscopy at 3 Å resolution. The gene therapy vector, AAV-DJ, is a hybrid of natural serotypes that was previously derived by directed evolution, selecting for hepatocyte entry and resistance to neutralization by human serum. The structure of AAV-DJ differs from that of parental serotypes in two regions where neutralizing antibodies bind, so immune escape appears to have been the primary driver of AAV-DJ's directed evolution. Fondaparinux is an analog of cell surface heparan sulfate to which several AAVs bind during entry. Fondaparinux interacts with viral arginines at a known heparin binding site, without the large conformational changes whose presence was controversial in low-resolution imaging of AAV2-heparin complexes. The glycan density suggests multi-modal binding that could accommodate sequence variation and multivalent binding along a glycan polymer, consistent with a role in attachment, prior to more specific interactions with a receptor protein mediating entry.
已通过冷冻电子显微镜在3埃分辨率下确定了腺相关病毒（AAV）-DJ单独以及与磺达肝癸钠复合时的原子结构。基因治疗载体AAV-DJ是天然血清型的杂交体，它先前是通过定向进化获得的，选择用于肝细胞进入和对人血清中和的抗性。AAV-DJ的结构在中和抗体结合的两个区域与亲本血清型不同，因此免疫逃逸似乎是AAV-DJ定向进化的主要驱动力。磺达肝癸钠是细胞表面硫酸乙酰肝素的类似物，几种AAV在进入过程中会与之结合。磺达肝癸钠在已知的肝素结合位点与病毒精氨酸相互作用，没有在AAV2-肝素复合物的低分辨率成像中存在争议的大的构象变化。聚糖密度表明多模式结合，这可以适应沿着聚糖聚合物的序列变异和多价结合，这与在与介导进入的受体蛋白进行更特异性相互作用之前在附着中的作用一致。
