纤溶酶原Ala601Thr突变导致的遗传性异常纤溶酶原血症表型与基因型分析  

Phenotype and genotype analysis of in herited dysplasminogenemia caused by plasminogen Ala601Thr mutation

作　　者：程晓丽[1] 杨丽红[1] 黄国咏 朱丽青[1] 金艳慧[1] 王明山[1] 

机构地区：[1]温州医科大学附属第一医院医学检验中心,325000

出　　处：《中华检验医学杂志》2016年第5期366-371,共6页Chinese Journal of Laboratory Medicine

基　　金：浙江省自然科学基金青年基金（Q15H200007）;国家自然科学基金（8150080275）

摘　　要：目的对1个遗传性异常纤溶酶原血症家系进行表型及基因突变检测，寻找其致病基因。方法2015年5月10日温州医科大学附属第一医院神经内科门诊，收治1例因“半月前无明显诱因出现头痛，两侧颞部胀痛，伴呕吐，复视，睡眠差”的25岁男性患者，入院诊断“上矢状窦静脉血栓形成”。采集先证者及其家系成员（共3代14人）外周静脉血标本，在全自动血凝仪上检测其凝血酶原时间（PT）、活化部分凝血活酶时间（Avrr）、凝血酶时间（TT）、纤维蛋白原（FIB）、D-二聚体（D．D）及纤维蛋白原降解产物（FDP）；采用发色底物法和免疫火箭电泳法分别检测纤溶酶原活性（PLG：A）和纤溶酶原抗原（PLG：Ag）。PCR扩增纤溶酶原（PLG）基因19个外显子及其5’和3’侧翼区。用DNA直接测序法分析扩增产物，并通过反向测序验证突变位点。采用生物信息学预测软件（SIFT，PolyPhen-2和MutationTaster）分析突变对蛋白质功能的影响。采用Swiss—PdbViewer软件对突变位点进行模型分析。结果先证者及6位家庭成员PLG：A降低约50％，PLG：Ag正常。先证者及其奶奶、父亲的D—D和FDP值轻度升高。DNA测序结果表明，先证者及这6位成员PLG基因15号外显子g．38829G〉A杂合错义突变，导致p．Ala601Thr。生物信息学软件预测结果表明此突变可影响蛋白质的功能。蛋白质模型分析显示突变改变了氨基酸之间的疏水作用力和氢键，从而影响了蛋白质的稳定性。且所有家系成员5’非翻译区均存在g．2501C〉A杂合SNP。结论在该遗传性异常纤溶酶原血症家系中发现的PLG基因P．Ala601Thr杂合突变与纤溶酶原活性水平降低有关。异常纤溶酶原血症及该突变位点均为国内首次报道。Objective To detectthe phenotype and gene mutations underlying aninherited dysplasminogenemia pedigree and search the virulence gene. Methods The peripheral venous blood samples of the proband and his family members （ fourteen subjects of three generations in total） were collected, and their prothrombin time（PT） , activated partial thromboplastin time（APTT） , thrombin time（TT） , fibrinogen （FIB）, fibrinogen degradation products （FDP）, D-dimmer （D-D）weretested on a STAGO analyzer, the plasminogen activity（ PLG ： A） and plasminogen antigen（ PLG ： Ag） were analyzedby thechromogenic substrate assay and rocket immunoelectrophoresis, respectively. All 19 exons, 5′ and 3′ untranslated regions of PLG were amplified with PCR. Direct DNA sequencing was used to analyze the amplified products, which were confirmed by backward sequencing. Three bioinformatics online softwares （SIFT, PolyPhen-2 and MutationTaster） were used to forecast the possible impact of the mutations on the protein function. At last ,the model analysis of mutate site was taken on a Swiss-Pdb Viewer software. Results The PLG ： Avalue of the proband and other 6 family members were decreased to the half, while the PLG：Ag was normal. The D-D and FDP value of the proband, his grandma and father were slightly higher. DNA sequencing has revealed that the proband and the other 6 members of this family had the same mutation of g. 38829G 〉 A in exon 15, leading to the missense mutationp. Ala601Thr. The results of bioinformatics softwares showed that the mutation could affect the thePLGfunction. Protein model analysis indicated that the hydrophobic interaction force and hydrogen bond between the amino acids were changed, which might affect the stability of the PLG. In addition, all the members of this family take the heterozygous SNP of g. 2501C 〉 A in the 5 UTR. Conclusions The p. Ala601Thr found in the inherited dysplasminogenemia pedigree in the exon 15 was responsible for the reduced PLG： A of the family,

关 键 词：突变 误义 纤溶蛋白溶酶原 多态性 单核苷酸 

分 类 号：R446.1[医药卫生—诊断学] R596.1[医药卫生—临床医学]