Department of Applied Science and Technology, Politecnico di Torino, Torino, Italy.
National Institute for Biological Standards and Control, Hertfordshire, United Kingdom.
PLoS One. 2022 Feb 7;17(2):e0262409. doi: 10.1371/journal.pone.0262409. eCollection 2022.
Allosteric disulfide bonds permit highly responsive, transient 'switch-like' properties that are ideal for processes like coagulation and inflammation that require rapid and localised responses to damage or injury. Haemophilia A (HA) is a rare bleeding disorder managed with exogenous coagulation factor(F) VIII products. FVIII has eight disulfide bonds and is known to be redox labile, but it is not known how reduction/oxidation affects the structure-function relationship, or its immunogenicity-a serious complication for 30% severe HA patients. Understanding how redox-mediated changes influence FVIII can inform molecular engineering strategies aimed at improving activity and stability, and reducing immunogenicity. FVIII is a challenging molecule to work with owing to its poor expression and instability so, in a proof-of-concept study, we used molecular dynamics (MD) to identify which disulfide bonds were most likely to be reduced and how this would affect structure/function; results were then experimentally verified. MD identified Cys1899-Cys1903 disulfide as the most likely to undergo reduction based on energy and proximity criteria. Further MD suggested this reduction led to a more open conformation. Here we present our findings and highlight the value of MD approaches.
变构二硫键允许高度响应、瞬态“开关样”特性，非常适合需要对损伤或伤害做出快速和局部反应的过程，如凝血和炎症。血友病 A (HA) 是一种罕见的出血性疾病，用外源性凝血因子 (FVIII) 产品进行管理。FVIII 有 8 个二硫键，已知其具有氧化还原不稳定性，但目前尚不清楚还原/氧化如何影响结构-功能关系或其免疫原性——这是 30%严重 HA 患者的一个严重并发症。了解氧化还原介导的变化如何影响 FVIII 可以为旨在提高活性和稳定性以及降低免疫原性的分子工程策略提供信息。FVIII 是一种具有挑战性的分子，因为其表达不佳且不稳定，因此，在一项概念验证研究中，我们使用分子动力学 (MD) 来确定哪些二硫键最有可能被还原，以及这将如何影响结构/功能；然后通过实验验证了结果。MD 根据能量和接近度标准确定 Cys1899-Cys1903 二硫键最有可能发生还原。进一步的 MD 表明，这种还原导致更开放的构象。在这里，我们提出了我们的发现，并强调了 MD 方法的价值。
