Effect of amino acid mutations on the conformational dynamics of amyloidogenic immunoglobulin light-chains: A combined NMR and in silico study.
Effect of amino acid mutations on the conformational dynamics of amyloidogenic immunoglobulin light-chains: A combined NMR and in silico study.
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DOI：
10.1038/s41598-017-10906-w
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发表时间：
2017-09-04
影响因子：
4.6
通讯作者：
Jaroniec CP
中科院分区：
文献类型：
--
作者：
Mukherjee S;Pondaven SP;Hand K;Madine J;Jaroniec CP
应用15 N弛豫分散核磁共振波谱研究了与AL淀粉样变性相关的致病性κ4人免疫球蛋白轻链可变区SMA的构象动力学。与同源轻链LEN相比，LEN与SMA在8个位置不同，但在体内是非淀粉样蛋白生成的，我们发现SMA中的多个残基聚集在N-末端和CDR环周围，经历了由毫秒时间尺度蛋白质运动引起的相当大的构象交换加宽，与不稳定的二聚体界面一致。为了评估每个氨基酸取代对塑造SMA的动态构象景观的贡献，对LEN的每个SMA样点突变体进行NMR研究，然后对这些蛋白质的子集进行计算机模拟分析。这些研究表明，位于二聚体界面处的CDR 3环内或直接邻近CDR 3环的仅三个突变的组合，其显著地包括不稳定（Q89 H和Y 96 Q）和稳定（T94 H）突变，在很大程度上解释了LEN和SMA之间构象柔性的差异。总的来说，我们的研究表明，稳定和不稳定的突变的正确组合是免疫球蛋白轻链填充未折叠的中间体，导致淀粉样蛋白形成的关键，并强调轻链构象灵活性，热力学稳定性和淀粉样蛋白形成之间的相关性的复杂性。
The conformational dynamics of a pathogenic κ4 human immunoglobulin light-chain variable domain, SMA, associated with AL amyloidosis, were investigated by 15N relaxation dispersion NMR spectroscopy. Compared to a homologous light-chain, LEN, which differs from SMA at eight positions but is non-amyloidogenic in vivo, we find that multiple residues in SMA clustered around the N-terminus and CDR loops experience considerable conformational exchange broadening caused by millisecond timescale protein motions, consistent with a destabilized dimer interface. To evaluate the contribution of each amino acid substitution to shaping the dynamic conformational landscape of SMA, NMR studies were performed for each SMA-like point mutant of LEN followed by in silico analysis for a subset of these proteins. These studies show that a combination of only three mutations located within or directly adjacent to CDR3 loop at the dimer interface, which remarkably include both destabilizing (Q89H and Y96Q) and stabilizing (T94H) mutations, largely accounts for the differences in conformational flexibility between LEN and SMA. Collectively, our studies indicate that a correct combination of stabilizing and destabilizing mutations is key for immunoglobulin light-chains populating unfolded intermediates that result in amyloid formation, and underscore the complex nature of correlations between light-chain conformational flexibility, thermodynamic stability and amyloidogenicity.
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