一个新的ABO*A等位基因导致的AwB亚型及其分子机制研究

doi:10.16150/j.1671-2870.2021.06.006

摘要/Abstract

摘要：

目的: 对1例血型A_wB亚型个体进行血清学鉴定和基因检测,探讨血型A_wB亚型发生的分子学机制。方法: 使用标准血清学实验方法对1例血型A_wB亚型个体进行ABO血清学定型,并对其ABO基因7个外显子及其侧翼序列进行PCR扩增、基因克隆和测序分析;采用Chimera软件进行ABO基因编码糖基转移A酶（glycosyl transferase A,GTA）突变体构建,用PyMOL软件作图并进行分析。结果: 血清学鉴定结果显示该个体为A_wB亚型。DNA克隆和基因测序分析结果显示,先证者的A基因为一个A1.02与A3.03顺式表达的等位基因,基因型为ABO*Avar/B.01。在国际输血协会数据库中查询,发现该变异A等位基应为一种新等位基因,其存在c.467C>T和c.838C>T错义突变,导致GTA发生p.P156L和p.L280F氨基酸置换。对GTA空间结构进行分析后认为,导致该表型的p.L280F未导致GTA蛋白整体结构的改变,但改变了GTA 280位氨基酸与周围氨基酸残基的氢键网络,导致局部构象改变。结论: 本研究发现了1个新的A等位基因导致的A_wB亚型,其形成机制为A1.02等位基因编码的糖基转移酶存在p.L280F突变,可能改变了邻近氨基酸间的作用力,从而导致A酶活性减弱,在与B等位基因共表达时形成A_wB亚型。

关键词: ABO血型系统, A_wB亚型, 基因突变

Abstract:

Objective: To investigate the molecular mechanism of A_wB subtype by performing serological and gene detection in a Chinese individual. Methods: One case of A_wB subtype was determined by ABO serology using standard serological method, and 7 exons of ABO gene and their flanks were amplified by PCR, and then cloned and sequenced. Glycosyl Transferase A (GTA) mutants were constructed by Chimera software, and was mapped and analyzed by PyMOL software. Results: Serological identification showed that this individual had A_wB subtype. DNA cloning and gene sequen-cing analysis showed that the A gene from the proband was A cis-expressed allele of A1.02 and A3.03, and the genotype was ABO*Avar/B.01. In the ISBT database, the A allele of this variant should be a novel allele, with missense mutations in c. 467C>T and c. 838C>T, which could result in p.P.156L and p.L.280F amino acid replacement in GTA. Analysis of the spatial structure of GTA showed, p.L280F resulted in the phenotypes, and did not lead to changes in the overall structure of GTA proteins, which altered the hydrogen bond network between amino acid 280 and surrounding amino acid residues, resulting in local conformation changes. Conclusions: A_wB subtype caused by a novel A allele-is reported for the first time. The formation mechanism may be as follows: p.L280F mutations in glycosyltransferases encoded by the A1.02 allele alter the interactions between adjacent amino acids, resulting in reduced activity of the A enzyme,and A_wB subtype occurs when co-expressing with the B allele.

Key words: ABO blood group system, A_wB subtype, Genetic mutation

中图分类号:

R446.11⁺2

周璐, 雷航, 洪叶, 金爽, 董永勤, 王学锋, 蔡晓红. 一个新的ABO*A等位基因导致的A_wB亚型及其分子机制研究[J]. 诊断学理论与实践, 2021, 20(06): 547-551.

ZHOU Lu, LEI Hang, HONG Ye, JIN Shuang, DONG Yongqin, WANG Xuefeng, CAI Xiaohong. A_wB subtype caused by a novel ABO *A allele and its molecular mechanisms[J]. Journal of Diagnostics Concepts & Practice, 2021, 20(06): 547-551.

图/表 2

参考文献 25

[1]	Daniels G. Human blood groups[M]. 3rd ed. Oxford: Blackwell,Science, 2013.
[2]	Fung MK, Eder AF, Spitalnik S, et al. AABB technical manual[M]. 19th ed. Bethesda(MD): American Associa tion of Blood Banks, 2017:785-786.
[3]	Cai XH, Jin S, Liu X, et al. Molecular genetic analysis for the B subgroup revealing two novel alleles in the ABO gene[J]. Transfusion, 2008, 48(11):2442-2447. doi: 10.1111/j.1537-2995.2008.01878.x URL
[4]	Pettersen EF, Goddard TD, Huang CC, et al. UCSF Chimera--a visualization system for exploratory research and analysis[J]. J Comput Chem, 2004, 25(13):1605-1612. doi: 10.1002/jcc.20084 pmid: 15264254
[5]	Ogasawara K, Yabe R, Uchikawa M, et al. Recombination and gene conversion-like events may contribute to ABO gene diversity causing various phenotypes[J]. Immunogenetics, 2001, 53(3):190-199. pmid: 11398963
[6]	曹红荣, 孟宪军, 王照军, 等. ABO亚型鉴定结果分析与报告——附B(A)04亚型1例[J]. 临床血液学杂志, 2017, 30(2):152-154.
[7]	杨璇, 郭思维, 洪强. A亚型患者血型鉴定流程及输血策略研究[J]. 中国卫生标准管理, 2019, 10(23):131-134.
[8]	井忠翠, 孙波, 孟子凡, 等. 血型基因检测在ABO和RhD血型鉴定困难患者输血中的应用价值[J]. 精准医学杂志, 2019, 34(4):337-340.
[9]	林晨蓓, 王秀娣, 朱碎永, 等. A3亚型血型的血清学和分子生物学鉴定分析[J]. 中国卫生检验杂志, 2018, 28(7):801-802,829.
[10]	段莉, 王远杰, 梁春艳, 等. ABO疑难血型的血清学检测及配血处理[J]. 四川医学, 2017, 38(5):489-491.
[11]	王天菊, 左琴琴, 齐珺, 等. O新等位基因与B抗原弱表达的研究[J]. 中国输血杂志, 2017, 30(10):1149-1152.
[12]	陈萍, 沈雨青, 张水木. 类孟买血型的鉴定及输血对策探讨[J]. 临床血液学杂志(输血与检验版), 2016, 29(3):508-509.
[13]	向东, 郭忠慧, 刘曦, 等. AB亚型细胞上H抗原强度的探讨[J]. 中国输血杂志, 2005, 18(3):189-191.
[14]	Sun CF, Yu LC, Chen IP, et al. Molecular genetic analysis for the A_e1 and A₃ alleles[J]. Transfusion, 2003, 43(8):1138-1144. doi: 10.1046/j.1537-2995.2003.00500.x URL
[15]	Pipatvanichkul A, Permpikul P, Vejbaesya S, et al. Mutation of ABO gene in Thai blood donors with A₃ phenotype[J]. J Med Assoc Thai, 2011, 94(3):379-385. pmid: 21560847
[16]	Chen YJ, Chen PS, Liu HM, et al. Novel polymorphisms in exons 6 and 7 of A/B alleles detected by polymerase chain reaction-single strand conformation polymorphism[J]. Vox Sang, 2006, 90(2):119-127. doi: 10.1111/j.1423-0410.2005.00729.x pmid: 16430670
[17]	Hosseini-Maaf B, Letts JA, Persson M, et al. Structural basis for red cell phenotypic changes in newly identified, naturally occurring subgroup mutants of the human blood group B glycosyltransferase[J]. Transfusion, 2007, 47(5):864-875. doi: 10.1111/j.1537-2995.2007.01203.x pmid: 17465952
[18]	Cai X, Jin S, Liu X, et al. Molecular genetic analysis of ABO blood group variations reveals 29 novel ABO subgroup alleles[J]. Transfusion, 2013, 53(11 Suppl 2):2910-2916.
[19]	金沙, 蔡晓红, 刘曦, 等. 上海地区献血人群cisAB和B(A)血型的研究[J]. 中国输血杂志, 2013, 26(12):1198-1201.
[20]	李茵, 赵倩, 苏蔓, 等. 罕见的A₃亚型的血清学和分子生物学研究[J]. 临床血液学杂志, 2021, 34(2):109-111.
[21]	韩斌, 冯智慧, 郭庆明, 等. 罕见的2例A307亚型的分子生物学研究[J]. 中国实验血液学杂志, 2016, 24(3):918-920.
[22]	喻琼, 梁延连, 邓志辉, 等. A₃等位基因的分子遗传分析[J]. 中国实验血液学杂志, 2005, 13(1):135-139.
[23]	Zhu F, Tao S, Xu X, et al. Distribution of ABO blood group allele and identification of three novel alleles in the Chinese Han population[J]. Vox Sang, 2010, 98(4):554-559. doi: 10.1111/j.1423-0410.2009.01291.x pmid: 20003128
[24]	Yamamoto F, McNeill PD, Hakomori S. Human histo-blood group A2 transferase coded by A2 allele, one of the A subtypes, is characterized by a single base deletion in the coding sequence, which results in an additional domain at the carboxyl terminal[J]. Biochem Biophys Res Commun, 1992, 187(1):366-374. doi: 10.1016/S0006-291X(05)81502-5 URL
[25]	陈萍, 张水木, 雷航, 等. 血清学鉴定与基因检测互补验证 ABO 疑难血型的应用探讨[J]. 诊断学理论与实践, 2020, 19(1):69-73.