一例遗传性凝血因子F13缺陷症家系的基因变异分析

Analysis of genetic variant in a Chinese pedigree with hereditary factor XIII deficiency

目的分析一个遗传性凝血因子XIII(coagulation factor XIII,FXIII)缺陷症患者一种新的基因变异,初步探讨其分子致病机制。方法PCR扩增F13A1、F13B基因所有外显子、侧翼序列以及5′端、3′端非翻译区,DNA直接测序。采用ClustalX-2.1-win软件分析氨基酸变异位点的保守性;用Mutation Taster、PolyPhen-2、PROVEAN、SIFT 4个在线生物信息学软件分析变异位点对蛋白功能的影响;用Swiss-PdbViewer软件对变异位点进行蛋白模型和氨基酸相互作用分析。结果在FXIII缺陷症患者F13A1基因第4外显子发现c.515G>C(p.Arg171Pro)杂合错义变异,该变异在同源物种间高度保守,4个在线生物信息学软件均显示p.Arg171Pro变异可能影响FXIII蛋白功能。蛋白模型分析显示,野生型Arg171与Pro27、Thr28各有1个氢键,与Glu102有2个氢键;当发生p.Arg171Pro变异后,Arg171与Pro27、Glu102之间的3个氢键均消失,形成一苯环结构,使蛋白质的内部结构发生了改变。结论F13B基因未发现变异。F13A1基因p.Arg171Pro杂合错义变异可能与该患者FXIII水平降低有关;p.Arg171Pro变异为国内外尚未报道过的新的F13A1基因变异。

Objective To explore the genetic basis for a patient with factor XIII(FXIII)deficiency.Methods All exons of the F13A1 and F13B genes were amplified by PCR and sequenced directly.The sequencing was performed with a reverse primer if a variant was found.Conservation of variant site was analyzed by the ClustalX software.Four online bioinformatic software including MutationTaster,PolyPhen-2,PROVEAN and SIFT were used to predict the function of the mutation site.The Swiss-PdbViewer software was applied to analyze the changes in the protein model and intermolecular force.Results The proband was found to harbor a novel c.515G>C(p.Arg171Pro)variant of the F13A1 gene.The corresponding amino acid Arg171 is conserved among homologous species.Bioinformatic analysis indicated that Arg171Pro variant may affect the protein function.Protein model analysis showed that in the wild-type,there is one hydrogen bond between Arg171 and Pro27;one hydrogen bond between Arg171 and Thr28;two hydrogen bonds between Arg171 and Glu102.When Arg171 was mutated to Pro171,the three hydrogen bonds between Arg171 and Pro27,Glu102 are all disappeared and formed a new benzene ring which might affect the stability of the protein structure.No variant was found in the F13B gene.Conclusion The Arg171Pro variant may account for the decreased FXIII level.Above finding has enriched the spectrum of F13A1 gene variants.