Al-Allaf Faisal A, Taher Mohiuddin M, Abduljaleel Zainularifeen, Bouazzaoui Abdellatif, Athar Mohammed, Bogari Neda M, Abalkhail Halah A, Owaidah Tarek Ma
Faculty of Medicine, Department of Medical Genetics, Umm Al-Qura University, Makkah, Kingdom of Saudi Arabia; Molecular Diagnostics Unit, Department of Laboratory Medicine and Blood Bank, King Abdullah Medical City, Makkah, Kingdom of Saudi Arabia; Science and Technology Unit, Umm Al-Qura University, Makkah, Kingdom of Saudi Arabia; These authors contributed equally to this study.
Faculty of Medicine, Department of Medical Genetics, Umm Al-Qura University, Makkah, Kingdom of Saudi Arabia; Science and Technology Unit, Umm Al-Qura University, Makkah, Kingdom of Saudi Arabia; These authors contributed equally to this study.
J Clin Med Res. 2017 Apr;9(4):317-331. doi: 10.14740/jocmr2876w. Epub 2017 Feb 21.
Hemophilias A and B are X-linked bleeding disorders caused by mutations in the factor VIII and factor IX genes, respectively. Our objective was to identify the spectrum of mutations of the factor VIII and factor IX genes in Saudi Arabian population and determine the genotype and phenotype correlations by molecular dynamics (MD) simulation.
For genotyping, blood samples from Saudi Arabian patients were collected, and the genomic DNA was amplified, and then sequenced by Sanger method. For molecular simulations, we have used softwares such as CHARMM (Chemistry at Harvard Macromolecular Mechanics; http://www.charmm-gui.org) and GROMACS. In addition, the secondary structure was determined based on the solvent accessibility for the confirmation of the protein stability at the site of mutation.
Six mutations (three novel and three known) were identified in factor VIII gene, and six mutations (one novel and five known) were identified in factor IX gene. The factor VIII novel mutations identified were c.99G>T, p. (W33C) in exon 1, c.2138 DelA, p. (N713Tfs*9) in eon14, also a novel mutation at splicing acceptor site of exon 23 c.6430 - 1G>A. In factor IX, we found a novel mutation c.855G>C, p. (E285D) in exon 8. These novel mutations were not reported in any factor VIII or factor IX databases previously. The deleterious effects of these novel mutations were confirmed by PolyPhen2 and SIFT programs.
The protein functional and structural studies and the models built in this work would be appropriate for predicting the effects of deleterious amino acid substitutions causing these genetic disorders. These findings are useful for genetic counseling in the case of consanguineous marriages which is more common in the Saudi Arabia.
血友病A和B是分别由凝血因子VIII和凝血因子IX基因突变引起的X连锁出血性疾病。我们的目的是确定沙特阿拉伯人群中凝血因子VIII和凝血因子IX基因的突变谱，并通过分子动力学（MD）模拟确定基因型和表型的相关性。
为了进行基因分型，收集了沙特阿拉伯患者的血液样本，扩增基因组DNA，然后采用桑格法进行测序。对于分子模拟，我们使用了诸如CHARMM（哈佛大分子力学化学；http://www.charmm-gui.org）和GROMACS等软件。此外，基于溶剂可及性确定二级结构，以确认突变位点处蛋白质的稳定性。
在凝血因子VIII基因中鉴定出6个突变（3个新突变和3个已知突变），在凝血因子IX基因中鉴定出6个突变（1个新突变和5个已知突变）。鉴定出的凝血因子VIII新突变是外显子1中的c.99G>T，p.（W33C），外显子14中的c.2138 DelA，p.（N713Tfs*9），以及外显子23剪接受体位点处的一个新突变c.6430 - 1G>A。在凝血因子IX中，我们在外显子8中发现了一个新突变c.855G>C，p.（E285D）。这些新突变以前在任何凝血因子VIII或凝血因子IX数据库中均未报道。这些新突变的有害影响通过PolyPhen2和SIFT程序得到证实。
本研究中进行的蛋白质功能和结构研究以及建立的模型将适用于预测导致这些遗传疾病的有害氨基酸替代的影响。这些发现对于沙特阿拉伯较为常见的近亲结婚情况下的遗传咨询很有用。
