Mukherjee Souhrid, Cassini Thomas A, Hu Ningning, Yang Tao, Li Bian, Shen Wangzhen, Moth Christopher W, Rinker David C, Sheehan Jonathan H, Cogan Joy D, Newman John H, Hamid Rizwan, Macdonald Robert L, Roden Dan M, Meiler Jens, Kuenze Georg, Phillips John A, Capra John A
Department of Biological Sciences, Vanderbilt University, Nashville, TN 37235, USA.
Center for Structural Biology, Vanderbilt University, Nashville, TN 37235, USA.
HGG Adv. 2022 Jul 19;3(4):100131. doi: 10.1016/j.xhgg.2022.100131. eCollection 2022 Oct 13.
Whole-exome sequencing (WES) in the clinic has identified several rare monogenic developmental and epileptic encephalopathies (DEE) caused by ion channel variants. However, WES often fails to provide actionable insight for rare diseases, such as DEEs, due to the challenges of interpreting variants of unknown significance (VUS). Here, we describe a "personalized structural biology" (PSB) approach that leverages recent innovations in the analysis of protein 3D structures to address this challenge. We illustrate this approach in an Undiagnosed Diseases Network (UDN) individual with DEE symptoms and a VUS in (p.V469L), the Kv3.2 voltage-gated potassium channel. A nearby variant (p.V471L) was recently suggested to cause DEE-like phenotypes. Computational structural modeling suggests that both affect protein function. However, despite their proximity, the p.V469L variant is likely to sterically block the channel pore, while the p.V471L variant is likely to stabilize the open state. Biochemical and electrophysiological analyses demonstrate heterogeneous loss-of-function and gain-of-function effects, as well as differential response to 4-aminopyridine treatment. Molecular dynamics simulations illustrate that the pore of the p.V469L variant is more constricted, increasing the energetic barrier for K permeation, whereas the p.V471L variant stabilizes the open conformation. Our results implicate variants in as causative for DEE and guide the interpretation of a UDN individual. They further delineate the molecular basis for the heterogeneous clinical phenotypes resulting from two proximal pathogenic variants. This demonstrates how the PSB approach can provide an analytical framework for individualized hypothesis-driven interpretation of protein-coding VUS.
临床全外显子组测序（WES）已鉴定出几种由离子通道变异引起的罕见单基因发育性和癫痫性脑病（DEE）。然而，由于解释意义未明变异（VUS）存在挑战，WES往往无法为诸如DEE等罕见疾病提供可采取行动的见解。在此，我们描述了一种“个性化结构生物学”（PSB）方法，该方法利用蛋白质三维结构分析方面的最新创新来应对这一挑战。我们在一名患有DEE症状且Kv3.2电压门控钾通道存在VUS（p.V469L）的未确诊疾病网络（UDN）个体中展示了这种方法。最近有研究表明，附近的一个变异（p.V471L）会导致类似DEE的表型。计算结构建模表明，这两种变异都会影响蛋白质功能。然而，尽管它们位置相近，但p.V469L变异可能在空间上阻塞通道孔，而p.V471L变异可能稳定开放状态。生化和电生理分析表明存在功能丧失和功能获得的异质性效应，以及对4 - 氨基吡啶治疗的不同反应。分子动力学模拟表明，p.V469L变异的孔更狭窄，增加了钾离子通透的能量屏障，而p.V471L变异稳定了开放构象。我们的结果表明该基因中的变异是DEE的病因，并指导了对一名UDN个体的解读。它们进一步阐明了由两个相邻致病变异导致的临床表型异质性的分子基础。这证明了PSB方法如何能够为蛋白质编码VUS的个体化假设驱动解读提供一个分析框架。
