Nila Nurun Nahar, Mahmud Zimam, Paul Anik, Rahman Taibur, Hossain Howlader Md Zakir, Hosen Md Ismail
Department of Biochemistry and Molecular Biology, University of Dhaka, Dhaka-1000, Bangladesh.
Heliyon. 2024 Jun 18;10(12):e33110. doi: 10.1016/j.heliyon.2024.e33110. eCollection 2024 Jun 30.
The Alternative Lengthening of Telomeres (ALT) pathway represents a non-canonical mechanism of telomere maintenance that operates independently of the conventional telomerase activity. The three biologically significant proteins, designated as SMARCAL1 (SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A-like protein 1), DAXX (Death domain-associated protein 6) and ATRX (alpha-thalassemia/mental retardation, X-linked) are associated with ALT in certain cancer types. The purpose of this study was to identify the most high-risk nsSNPs (non-synonymous Single Nucleotide Polymorphisms) within these three genes and assess their impacts on the structure and function of the proteins they encode.
The reported genetic polymorphisms of and genes were retrieved from the Ensembl database. Later, various computational tools like PROVEAN, PolyPhen2, SNPs and GO, SNAP2, Predict-SNP, Panther and PMut were used to predict the most deleterious nsSNPs. MutPred was used to understand the underlying molecular reasons of those nsSNPs being deleterious, followed by prediction of Post Translational Modification Sites (PTMs) using ModPred. I-Mutant and MUpro were used to predict the effect of SNP on energy stability. Later, 3D clustering analysis was done using Mutation 3D server. Moreover, ConSurf was utilized to identify the conservation scores of wild-type amino acids. Additionally, the NCBI conserved domain search tool was employed to pinpoint conserved domains within these three proteins. Project-Hope helped for biophysical validation, followed by prediction of these genes' interaction and function by using GeneMANIA.
Analysis on SMARCAL1 protein revealed that among 665 nsSNPs, four were identified as the most deleterious: L578S, T581S, P582A, and P582S. Similarly, within the DAXX protein, among a pool of 480 nsSNPs, P284S, R230C, and R230S were found out to be the most deleterious variants. In case of ATRX protein, V178D, R246C, and V277G, from the total of 1009 nsSNPs, were predicted to be the most deleterious. All these nsSNPs were found to occur at residue positions that are 100 % conserved within protein domains and were predicted to be most damaging from both structural and functional perspectives and highly destabilizing to their corresponding proteins.
Computational investigation on the 3 proteins-SMARCAL1, DAXX and ATRX through different bioinformatics analysis tools concludes that the identified high risk nsSNPs of these proteins are pathogenic SNPs. These variants potentially exert functional and structural influences, thus making them valuable candidates for future genetic studies.
端粒替代延长（ALT）途径代表了一种不依赖于传统端粒酶活性的非经典端粒维持机制。三种具有生物学意义的蛋白质，即SMARCAL1（SWI/SNF相关的基质相关肌动蛋白依赖性染色质亚家族A样蛋白1）、DAXX（死亡结构域相关蛋白6）和ATRX（α地中海贫血/智力发育迟缓，X连锁）在某些癌症类型中与ALT相关。本研究的目的是在这三个基因中鉴定出风险最高的非同义单核苷酸多态性（nsSNPs），并评估它们对所编码蛋白质的结构和功能的影响。
从Ensembl数据库中检索和基因的已报道遗传多态性。随后，使用多种计算工具，如PROVEAN、PolyPhen2、SNPs和GO、SNAP2、Predict-SNP、Panther和PMut来预测最有害的nsSNPs。使用MutPred来了解这些nsSNPs有害的潜在分子原因，随后使用ModPred预测翻译后修饰位点（PTMs）。使用I-Mutant和MUpro预测SNP对能量稳定性的影响。随后，使用Mutation 3D服务器进行三维聚类分析。此外，利用ConSurf确定野生型氨基酸的保守分数。此外，使用NCBI保守结构域搜索工具来确定这三种蛋白质中的保守结构域。Project-Hope有助于进行生物物理验证，随后使用GeneMANIA预测这些基因的相互作用和功能。
对SMARCAL1蛋白的分析表明，在665个nsSNPs中，有四个被确定为最有害的：L578S、T581S、P582A和P582S。同样，在DAXX蛋白中，在480个nsSNPs中，发现P284S、R230C和R230S是最有害的变体。就ATRX蛋白而言，在总共1009个nsSNPs中，V178D、R246C和V277G被预测为最有害的。所有这些nsSNPs都发生在蛋白质结构域内100%保守的残基位置，并且从结构和功能角度来看都被预测为最具破坏性，并且对其相应蛋白质具有高度的不稳定作用。
通过不同的生物信息学分析工具对SMARCAL1、DAXX和ATRX这三种蛋白质进行的计算研究得出结论，这些蛋白质中鉴定出的高风险nsSNPs是致病性SNP。这些变体可能会产生功能和结构影响，因此使其成为未来基因研究的有价值候选对象。
