Department of Physics and Astronomy, The University of British Columbia, Vancouver, British Columbia V6T 1Z1, Canada.
Djavad Mowafaghian Centre for Brain Health, The University of British Columbia, Vancouver, British Columbia V6T 1Z1, Canada.
ACS Chem Neurosci. 2023 Aug 2;14(15):2603-2617. doi: 10.1021/acschemneuro.3c00007. Epub 2023 Jul 17.
Tau pathology is associated with many neurodegenerative disorders, including Alzheimer's disease (AD), where the spatio-temporal pattern of tau neurofibrillary tangles strongly correlates with disease progression, which motivates therapeutics selective for misfolded tau. Here, we introduce a new avidity-enhanced, multi-epitope approach for protein-misfolding immunogen design, which is predicted to mimic the conformational state of an exposed epitope in toxic tau oligomers. A predicted oligomer-selective tau epitope KLDFK was scaffolded by designing a β-helix structure that incorporated multiple instances of the 16-residue tau fragment VKSEKLDFKDRVQSKI. Large-scale conformational ensemble analyses involving Jensen-Shannon Divergence and the embedding depth showed that the multi-epitope scaffolding approach, employed in designing the β-helix scaffold, was predicted to better discriminate toxic tau oligomers than other "monovalent" strategies utilizing a single instance of an epitope for vaccine immunogen design. Using Rosetta, 10,000 sequences were designed and screened for the linker portions of the β-helix scaffold, along with a C-terminal stabilizing α-helix that interacts with the linkers, to optimize the folded structure and stability of the scaffold. Structures were ranked by energy, and the lowest 1% (82 unique sequences) were verified using AlphaFold. Several selection criteria involving AlphaFold are implemented to obtain a lead-designed sequence. The structure was further predicted to have free energetic stability by using Hamiltonian replica exchange molecular dynamics (MD) simulations. The synthesized β-helix scaffold showed direct binding in surface plasmon resonance (SPR) experiments to several antibodies that were raised to the structured epitope using a designed cyclic peptide. Moreover, the strength of binding of these antibodies to in vitro tau oligomers correlated with the strength of binding to the β-helix construct, suggesting that the construct presents an oligomer-like conformation and may thus constitute an effective oligomer-selective immunogen.
tau 病理学与许多神经退行性疾病有关，包括阿尔茨海默病 (AD)，tau 神经原纤维缠结的时空模式与疾病进展密切相关，这促使人们开发针对错误折叠 tau 的治疗方法。在这里，我们引入了一种新的亲和力增强的多表位方法，用于蛋白质错误折叠免疫原设计，该方法预计可模拟暴露于毒性 tau 寡聚物中的表位的构象状态。通过设计一个包含多个 16 残基 tau 片段 VKSEKLDFKDRVQSKI 的β-螺旋结构，支架了一个预测的寡聚体选择性 tau 表位 KLDFK。涉及 Jensen-Shannon 散度和嵌入深度的大规模构象整体分析表明，在设计β-螺旋支架时使用的多表位支架方法，预计比其他“单价”策略更好地区分毒性 tau 寡聚物，这些策略用于疫苗免疫原设计的单个表位。使用 Rosetta，设计并筛选了 10000 个序列，用于β-螺旋支架的连接部分，以及与连接子相互作用的 C 末端稳定的α-螺旋，以优化支架的折叠结构和稳定性。根据能量对结构进行排序，并使用 AlphaFold 验证最低的 1%（82 个独特序列）。实施了几个涉及 AlphaFold 的选择标准来获得主导设计序列。该结构进一步通过使用哈密顿复制交换分子动力学 (MD) 模拟来预测其自由能稳定性。在表面等离子体共振 (SPR) 实验中，合成的β-螺旋支架直接与针对该结构表位设计的环状肽产生的几种抗体结合。此外，这些抗体与体外 tau 寡聚物的结合强度与与β-螺旋构建体的结合强度相关，这表明该构建体呈现出类似寡聚物的构象，因此可能构成有效的寡聚体选择性免疫原。
