[Phenotype and genetic mutation analysis of an inherited protein C deficiency pedigree].

Objective: A phenotypic and gene mutation study was carried out to investigate the molecular mechanism of inherited protein C deficiency in a family with the disorder. Methods: The proband was a 21-year-old male who was admitted to hospital due to swelling of the left lower limb for 3 months and hemoptysis with chest tightness for more than 1 week. The clinical diagnosis was pulmonary embolism and deep vein thrombosis of the left lower limb. Plasma protein C activity, protein S activity and antithrombin Ⅲ activity were detected in the patient and their family members. Whole exon sequencing was used to analyze a total of 199 genes associated with thrombus susceptibility of the patient. After the mutation was found and Sanger sequencing was used to verify whether the family members carried the same gene mutations as the patient. The conservation of amino acid mutation sites was analyzed by using the software ClustalX-2.1-win. The damage of mutations to protein function was analyzed by PROVEAN and PolyPhen-2 online bioinformatics software. Finally, PyMOL 2.3 software was used to analyze the protein model. Results: The patient and four family members all had the identical heterozygous missense mutation c.1019 C>T (p. Thr340Met) in exon 9 of the protein C gene, resulting in various degrees of protein C deficiency. The Thr340 amino acid was discovered to be poorly conserved in seven homologous species after investigation with the clustalx-2.1-win software. P. Thr340Met was found to be a detrimental mutation by both PROVEAN and PolyPhen-2 online bioinformatics program. The mutation of Thr340 to Met340 caused the hydrogen bond between Thr340 and Gln226 to dissolve, changing the spatial arrangement of protein C, which might be the main explanation for the lower protein C activity, according to the protein model. Conclusions: Protein C deficiency in this family was caused by a hybrid missense mutation C. 1019 C>T (p. Thr340Met). Protein C deficiency may present in varying severity among mutation carriers at the same locus of the protein C gene. Whole-exome sequencing may be considered in young patients with spontaneous venous thromboembolism, even if there is no relevant family history.目的： 对一个遗传性蛋白C缺陷症家系进行表型和基因突变分析，探讨其发病的分子机制。 方法： 先证者为21岁男性，因“左下肢肿胀3个月余、咯血伴胸闷1周余”入院。临床诊断：肺栓塞，左下肢深静脉血栓形成。对患者及其家族成员进行血浆蛋白C活性、蛋白S活性、抗凝血酶Ⅲ活性检测；采用全外显子测序分析患者的 199 个血栓易感相关基因，发现突变后通过Sanger测序验证家族成员是否携带和患者相同位点的基因突变；再用ClustalX-2.1-win软件分析氨基酸突变位点的保守性；借助于PROVEAN和PolyPhen-2 在线生物信息学软件分析突变对蛋白质功能的危害程度；最后采用PyMOL 2.3软件进行蛋白质模型分析。 结果： 患者及4位家族成员蛋白C基因第9号外显子均携带同一个杂合错义突变c.1019C>T（p.Thr340Met），却表现为不同程度的蛋白C缺陷；经过ClustalX-2.1-win软件分析后发现，氨基酸Thr340在7种同源物种中不是高度保守；PROVEAN和PolyPhen-2在线生物信息学软件分析后均提示p.Thr340Met为有害突变；蛋白质模型分析提示：Thr340突变为Met340时，导致Thr340与Gln226之间的氢键断裂，改变了氨基酸的空间构型，这可能是蛋白C活性下降的主要原因。 结论： 蛋白C基因第9号外显子杂合错义突变c.1019C>T（p.Thr340Met）是导致该家族蛋白C缺陷症的主要原因。蛋白C基因同一位点突变的携带者可能表现为不同程度的蛋白C缺陷症。对于无诱因的静脉血栓栓塞年轻患者，即使没有相关的家族史，也可考虑行全外显子测序以明确病因。.