CAJ | 학술논문

目的：克隆、表达常见的人呼吸道腺病毒的主要中和抗原六邻体蛋白，分析重组表达产物的抗原性和免疫原性。方法分别PCR扩增克隆人3型、4型、7型腺病毒完整的六邻体蛋白基因，测序和序列比对并与GenBank上人腺病毒的六邻体蛋白进行同源性比对；对其主要的抗原区进行克隆、表达和纯化，表达产物免疫动物，ELISA和Western blot实验对表达产物的抗原性和免疫原性及其交叉反应进行分析。结果国内首次报道1株人4型腺病毒的六邻体基因序列，与GenBank中的NHRC3分离株核酸及氨基酸的同源性均大于99％；3种腺病毒包含全部高变区的六邻体部分区段在大肠杆菌中成功表达并纯化，重组蛋白可以被不同型别的人腺病毒免疫血清识别，重组蛋白免疫小鼠血清可以识别不同型别的人腺病毒，而型间反应有显著差异，预测了3种腺病毒六邻体型、型间特异性表位。结论成功表达人3型、4型、7型腺病毒六邻体蛋白主要抗原区，重组表达产物有强免疫原性，包含型间、型特异性抗原表位。
목적：극륭、표체상견적인호흡도선병독적주요중화항원륙린체단백，분석중조표체산물적항원성화면역원성。방법분별PCR확증극륭인3형、4형、7형선병독완정적륙린체단백기인，측서화서렬비대병여GenBank상인선병독적륙린체단백진행동원성비대；대기주요적항원구진행극륭、표체화순화，표체산물면역동물，ELISA화Western blot실험대표체산물적항원성화면역원성급기교차반응진행분석。결과국내수차보도1주인4형선병독적륙린체기인서렬，여GenBank중적NHRC3분리주핵산급안기산적동원성균대우99％；3충선병독포함전부고변구적륙린체부분구단재대장간균중성공표체병순화，중조단백가이피불동형별적인선병독면역혈청식별，중조단백면역소서혈청가이식별불동형별적인선병독，이형간반응유현저차이，예측료3충선병독륙린체형、형간특이성표위。결론성공표체인3형、4형、7형선병독륙린체단백주요항원구，중조표체산물유강면역원성，포함형간、형특이성항원표위。
Objective To clone and express the hexon protein of three prevalent human adenovi -rus strains causing respiratory disease and analyze the antigenic characteristics of the recombinant proteins . Methods The full length genes encoding hexon protein of human adenovirus serotype 3(HAdV3), serotype 4(HAdV4) and serotype 7(HAdV7) were cloned by PCR and sequenced , respectively.The alignment anal-ysis was performed by using hexon gene sequences from GenBank .The major antigenic regions of hexon pro-tein of the three serotypes were expressed in E.coli and purified.The antigenicity, immunogenicity and cross reactivity of the recombinant proteins were determined by ELISA and Western blot assay .Results The full length gene sequence encoding hexon protein of human adenovirus serotype 4 was firstly reported in China , which showed more than 99%homology in both nucleotide and amino acid sequences with the human adeno-virus type 4 NHRC3 strain.The partial hexon protein sequence of HAdV 3, HAdV4 and HAdV7 containing all of the 7 hyperviriable regions ( HVRs) were expressed in E.coli, respectively .The purified recombinant proteins could be recognized by antiserum of the three serotypes of adenovirus .The antiserum samples against the three recombinant proteins could cross-react with particles of the three serotypes of adenovirus . The possible type-and species-specific epitopes were predicted .Conclusion The major antigenic regions of hexon protein of the three serotypes were successfully expressed .The purified recombinant proteins contai-ning both intertypes and type-specific epitopes showed a strong immunogenicity .