ABO基因c.398T>C新变异导致Bweak表型的分子特性研究  

Study of the molecular characteristics of a Bweak phenotype due to a novel c.398T>C variant of the ABO gene

作　　者：应燕玲[1] 洪小珍[1] 张晶晶 马开荣[1] 刘瑛[1] 许先国[1] 何吉[1] 朱发明[1] Ying Yanling;Hong Xiaozhen;Zhang Jingjing;Ma Kairong;Liu Ying;Xu Xianguo;He Ji;Zhu Faming(Blood Center of Zhejiang Province,Key Laboratory of Blood Safety Research of Zhejiang Province,Hangzhou,Zhejiang 310052,China)

机构地区：[1]浙江省血液中心浙江省血液安全研究重点实验室,杭州310052

出　　处：《中华医学遗传学杂志》2023年第1期110-113,共4页Chinese Journal of Medical Genetics

基　　金：国家自然科学基金(81902137、82070195);浙江医药卫生科技计划(WKJ-ZJ-192、2019KY370)。

摘　　要：目的探讨1例Bweak亚型个体的分子机制。方法选取2016年12月5日于浙江省血液中心献血的1例受试者为研究对象。利用血清学方法鉴定受试者的ABO表型,用体外酶活性试验测定其血清中B糖基转移酶(GTB)的活性。用PCR扩增ABO基因第5~7外显子及侧翼序列并确定其基因型,采用T-A克隆技术分离单倍体并进行测序验证。用ProtParam和PSIPRED软件分析蛋白的一级理化性质和二级结构。用PolyPhen-2、SIFT、PROVEAN三种软件分析错义变异对蛋白的作用效应。结果受试者血清学检测为Bweak亚型,血清中存在抗B抗体。体外酶活性试验显示其GTB活性显著降低。单倍体克隆测序分析发现B等位基因上存在c.398T>C错义变异,为一个新的B等位基因,可导致GTB第133位的苯丙氨酸替换为丝氨酸(p.Phe133Ser)。生物信息学分析提示上述替换对蛋白的一级和二级结构影响不明显,但变异蛋白的热力学能量增加6.07 kcal/mol,严重降低了热稳定性,生物信息学预测该变异对蛋白功能有害。结论新等位基因ABO*B.01-398C是引起Bweak亚型抗原弱表达的机制,生物信息学分析有助于评估其结构和功能的变化。Objective To explore the molecular mechanism for an individual with Bweak subtype.Methods Serological methods were used to identify the proband′s phenotype.In vitro enzyme activity test was used to determine the activity of B-glycosyltransferase(GTB)in her serum.The genotype was determined by PCR amplification and direct sequencing of exons 5 to 7 and flanking sequences of the ABO gene.T-A cloning technology was used to isolate the haploids.The primary physical and chemical properties and secondary structure of the protein were analyzed with the ProtParam and PSIPRED software.Three software,including PolyPhen-2,SIFT,and PROVEAN,was used to analyze the effect of missense variant on the protein.Results Serological results showed that the proband′s phenotype was Bweak subtype and anti-B antibodies presented in her serum.In vitro enzyme activity assay showed that the GTB activity of the subject was significantly reduced.Analysis of the haploid sequence revealed a c.398T>C missense variant on the B allele,which resulted in a novel B allele.The 398T>C variant has caused a p.Phe133S substitution at position 133 of the GTB protein.Based on bioinformatic analysis,the amino acid substitution had no obvious effect on the primary and secondary structure of the protein,but the thermodynamic energy of the variant protein has increased to 6.07 kcal/mol,which can severely reduce the protein stability.Meanwhile,bioinformatic analysis also predicted that the missense variant was harmful to the protein function.Conclusion The weak expression of the Bweak subtype may be attributed to the novel allele of ABO*B.01-398C.Bioinformatic analysis is helpful for predicting the changes in protein structure and function.

关 键 词：Bweak变异型 B糖基转移酶 氨基酸置换 生物信息学 

分 类 号：R394[医药卫生—医学遗传学]