不同拓扑结构区域的HBV突变体对乙肝表面抗原合成和分泌的影响  

Effect of HBV mutants in different topological regions on the synthesis and secretion of HBsAg

作　　者：何培 张佩欣 李永刚 王佳[1] 彭传梅 袁勇[1] 张振华[3] 冯磊[1] HE Pei;ZHANG Peixin;LI Yonggang;WANG Jia;PENG Chuanmei;YUAN Yong;ZHANG Zhenhua;FENG Lei(Yan'an Hospital Affiliated to Kunming Medical University,Kunming,Yunnan 650000,China;The First Affiliated Hospital of USTC,Hefei,Anhui 230001,China;The Second Hospital of Anhui Medical University,Hefei,Anhui 230001,China;不详)

机构地区：[1]昆明医科大学附属延安医院,云南昆明650000 [2]中国科学技术大学第一附属医院,安徽合肥230001 [3]安徽医科大学第二附属医院,安徽合肥230000

出　　处：《中国热带医学》2024年第12期1519-1523,共5页China Tropical Medicine

基　　金：云南省教育厅-青年人才基础研究计划项目(No.2024J0276)

摘　　要：目的 隐匿性乙型肝炎病毒(hepatitis B virus, HBV)突变株具有较强传播能力和危害性,探究HBsAg抗原变异、合成及分泌,将为解决临床隐匿性乙型肝炎诊疗问题提供有益的指导。方法 对隐匿性HBV感染(occult hepatitis B virus infection, OBI)患者HBV基因序列成功克隆乙肝表面抗原(hepatitis B surface antigen, HBsAg)单基因突变体和多基因突变体,使用AlphaFold2.3软件对野生型基突变体结构模型进行蛋白相似性及稳定性分析,并将野生型、单及多位点突变体转染至Huh7细胞,分析不同突变体的细胞内外合成和分泌情况。结果 单突变位点Y100L、Y100C及C139R与野生型相比的均方根偏差(root mean square deviation, RMSD)值较小,G145R与野生型比较的RMSD值较大。Y100L热能差(ΔΔG)最小,其次是Y100C、C139R及G145R。3种多位点突变体与野生型相比的RMSD值均小于1,T63I/Y100L/Q101K热能差(ΔΔG)最小,T125A/F158L/V224G及S113T/F220C次之。Y100C、C139R、G145R合成、分泌以及分泌效率较野生型差异均有统计学意义(P<0.05)。S113T/F220C细胞内外的合成、分泌及分泌效率与野生型相比,差异均有统计学意义(P<0.05);T63I/Y100L/Q101K细胞内合成及分泌效率与野生型相比,差异有统计学意义(P<0.05)。结论 内质网腔区部分单点突变体在肝细胞内合成和分泌HBsAg降低;内质网腔区和其他结构区域组合的部分多点突变体在肝细胞内合成HBsAg增加及分泌效率降低。Objective Because occult hepatitis B virus(HBV)variants are highly transmissible and harmful,the investigation of HBsAg antigenic mutation,synthesis,and secretion will provide useful guidance for addressing clinical diagnosis and treatment of occult hepatitis B infection.Methods The HBV gene sequences of OBI patients were successfully cloned to obtain HBsAg single and multi-locus mutants.The protein similarity and stability of the wild-type and mutant structural model were analyzed using AlphaFold2.3 software,and the wild-type,single and multi-locus mutants were transfected into Huh7 cells to analyze the intra-and extracellular synthesis and secretion of different mutants.Results The single mutant loci Y100L,Y100C,and C139R showed smaller root mean squared deviation(RMSD)values compared to the wild type,while G145R had a larger RMSD value.Moreover,the Y100L mutation exhibited the smallest thermos-energetic difference(ΔΔG),followed by Y100C,C139R,and G145R.The RMSD values of the three multi-locus mutants were less than 1 compared to the wild type,and the RMSD values of T63I/Y100L/Q101K were the smallest,followed by T125A/F158L/V224G and S113T/F220C.The synthesis,secretion,and secretion efficiency of Y100C,C139R,and G145R were significantly different from the wild-type(P<0.05);the intracellular synthesis and secretion efficiency of S113T/F220C were statistically different compared to the wild-type(P<0.05);the intracellular synthesis and secretion efficiency of T63I/Y100L/Q101K were statistically different compared to the wild type(P<0.05).Conclusion Some single-point mutants in the luminal region of the endoplasmic reticulum reduce the synthesis and secretion of HBsAg in hepatocytes.Certain multi-loci mutants combining the luminal region of the endoplasmic reticulum with other structural regions increase the synthesis of HBsAg in hepatocytes but reduce secretion efficiency.

关 键 词：隐匿性乙型肝炎 HBV突变体 乙肝表面抗原 

分 类 号：R512.62[医药卫生—内科学]