Expression in Escherichia coli of a cloned DNA sequence encoding the pre-S2 region of hepatitis B virus.
Expression in Escherichia coli of a cloned DNA sequence encoding the pre-S2 region of hepatitis B virus.
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编码乙型肝炎病毒前 S2 区的克隆 DNA 序列在大肠杆菌中的表达。
DOI：
10.1073/pnas.82.22.7540
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发表时间：
1985
影响因子：
11.1
通讯作者：
Acs,G
中科院分区：
文献类型：
--
作者：
Offensperger,W;Wahl,S;Neurath,AR;Price,P;Strick,N;Kent,SB;Christman,JK;Acs,G
编码人乙肝病毒包膜蛋白Pre-S2区(氨基酸120-174；血清型ayw)的DNA序列被插入到pSKS105的lacZ基因中，产生了重组的pWS3。经pWS3转化后分离到的大肠杆菌M182 Delta菌株的Lac+菌落(LacIOPZYA)产生一种Pre-S2肽-β-半乳糖苷酶融合蛋白。该融合蛋白占菌体总蛋白的3%，经p-aminophenyl-beta-D-thiogalactoside-Sepharose.亲和层析纯化，均一度达90%以上。它可与兔抗adw血清型Pre-S2区氨基酸第120-145位氨基酸的抗体或乙肝病毒抗体免疫共沉淀。前S完全阻断了任何一种抗体与前S2肽-β-半乳糖苷酶融合蛋白的结合。这些结果表明，融合蛋白上存在与人工合成的前S(12 0-14 5)和天然乙肝病毒颗粒前S2序列上的决定簇密切相关的抗原决定簇。因此，转化pWS3的细菌可以提供丰富的Pre-S2-β-半乳糖苷酶融合蛋白的来源，这可能被证明是一种有用的诊断试剂，具有适合于ELISA检测的标志酶活性，或者作为一种免疫原，可能有助于主动预防乙肝。
A DNA sequence encoding the entire pre-S2 region (amino acids 120-174; serotype ayw) of human hepatitis B virus envelope protein has been inserted into the lacZ gene of the plasmid pSKS105 yielding a recombinant, pWS3. Lac+ colonies of the Escherichia coli M182 delta (lacIOPZYA), isolated after transformation with pWS3, produced a pre-S2 peptide-beta-galactosidase fusion protein. This fusion protein, which comprised as much as 3% of the total bacterial protein, was purified to greater than 90% homogeneity by affinity chromatography on p-aminophenyl-beta-D-thiogalactoside-Sepharose. It is immunoprecipitable with rabbit antibodies to a synthetic peptide corresponding to amino acids 120-145 of the pre-S2 region of serotype adw [pre-S(120-145)] or with antibodies to hepatitis B virus. pre-S(120-145) completely blocked the binding of either antibody to the pre-S2 peptide-beta-galactosidase fusion protein. These results indicate that there are antigenic determinants on the fusion protein that are closely related to, if not identical to, determinants on synthetic pre-S(120-145) and on pre-S2 sequences of native hepatitis B virus particles. Thus, bacteria transformed with pWS3 can provide an abundant source of pre-S2-beta-galactosidase fusion protein, which may prove useful either as a diagnostic reagent possessing marker enzyme activity suitable for ELISA tests or as an immunogen with potential to contribute to active prophylaxis of hepatitis B.