Nonketotic hyperglycinemia: Functional assessment of missense variants in GLDC to understand phenotypes of the disease
Nonketotic hyperglycinemia: Functional assessment of missense variants in GLDC to understand phenotypes of the disease
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DOI：
10.1002/humu.23208
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发表时间：
2017-06-01
期刊：
影响因子：
3.9
通讯作者：
Rodriguez-Pombo, Pilar
中科院分区：
文献类型：
--
作者：
Bravo-Alonso, Irene;Navarrete, Rosa;Rodriguez-Pombo, Pilar
对基因组数据的快速分析为具有特定疾病的临床和生化特征的患者提供了有效的突变确认。这是非酮症高甘氨酸血症（NKH）的情况，NKH是一种孟德尔疾病，主要由于GLDC基因突变导致新生儿和早期婴儿癫痫发作。然而，理解在该基因中鉴定的错义变体的影响是基因组学应用于临床实践的主要挑战。在此，对在26名NKH患者的队列中鉴定的19种GLDC错义变体进行了全面的功能和结构分析。突变体cDNA构建体在COS 7细胞中表达，随后进行GCS P-蛋白的酶测定和Western印迹分析，以评估残余活性和突变体蛋白稳定性。还进行了基于GCS P-蛋白的三维结构的分子建模的结构分析。我们鉴定了产生减毒表型的亚型变体，其改善了疾病的预后。结构分析使我们能够解释突变对蛋白质稳定性和催化活性的影响，为临床结果和疾病严重程度提供分子证据。此外，我们确定了重要数量的突变体，其功能丧失与不稳定性相关，因此，是使用折叠治疗方法进行救援的潜在目标。
The rapid analysis of genomic data is providing effective mutational confirmation in patients with clinical and biochemical hallmarks of a specific disease. This is the case for nonketotic hyperglycinemia (NKH), a Mendelian disorder causing seizures in neonates and early-infants, primarily due to mutations in the GLDC gene. However, understanding the impact of missense variants identified in this gene is a major challenge for the application of genomics into clinical practice. Herein, a comprehensive functional and structural analysis of 19 GLDC missense variants identified in a cohort of 26 NKH patients was performed. Mutant cDNA constructs were expressed in COS7 cells followed by enzymatic assays and Western blot analysis of the GCS P-protein to assess the residual activity and mutant protein stability. Structural analysis, based on molecular modeling of the 3D structure of GCS P-protein, was also performed. We identify hypomorphic variants that produce attenuated phenotypes with improved prognosis of the disease. Structural analysis allows us to interpret the effects of mutations on protein stability and catalytic activity, providing molecular evidence for clinical outcome and disease severity. Moreover, we identify an important number of mutants whose loss-of-functionality is associated with instability and, thus, are potential targets for rescue using folding therapeutic approaches.