A new epitope tag from hepatitis B virus preS1 for immunodetection, localization and affinity purification of recombinant proteins

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dc.contributor.authorM S Oh-
dc.contributor.authorK S Kim-
dc.contributor.authorY K Jang-
dc.contributor.authorC Y Maeng-
dc.contributor.authorS H Min-
dc.contributor.authorM H Jang-
dc.contributor.authorS O Yoon-
dc.contributor.authorJ H Kim-
dc.contributor.authorHyo Jeong Hong-
dc.date.accessioned2017-04-19T09:00:40Z-
dc.date.available2017-04-19T09:00:40Z-
dc.date.issued2003-
dc.identifier.issn0022-1759-
dc.identifier.uri10.1016/j.jim.2003.08.010ko
dc.identifier.urihttps://oak.kribb.re.kr/handle/201005/6358-
dc.description.abstractPreviously, a murine monoclonal antibody (mAb) KR127 (IgG2a/κ) that binds specifically to the preS1 of hepatitis B virus (HBV) was generated and the fine epitope was mapped to amino acids (aa) 37-45 (NSNNPDWDF). In this current study, the epitope in combination with KR127 was tested for protein tagging. Initially, to evaluate the importance of each residue of the KR127 epitope in antibody binding, alanine substitution mutants of the epitope were constructed and characterized for KR127 binding by immunoblot analysis and competition ELISA. The results showed that substitution of Ser38 by alanine (S38A) increased the affinity to KR127. The mutated epitope (NANNPDWDF), designated S1 tag, was fused to the amino (N)- or carboxyl (C)-terminus of three human recombinant proteins, soluble B lymphocyte stimulator (sBLyS), the N-terminal domain of thrombopoietin (nTPO), and a mitochondrial ribosomal protein (CGI-113) for expression in mammalian cells, while it was fused to the N- or C-terminus of two proteins, a single-chain antibody fragment (ScFv) and the carboxyl-terminal domain (PAc) of the protective antigen of Bacillus anthracis for expression in Escherichia coli. The immunodetection, immunoprecipitation, and affinity purification of the expressed S1-tagged proteins by KR127 were successfully demonstrated. In addition, a KR127 mutant (AP2) with higher affinity, Kd (0.9 nM), for the S1 tag compared to that (20 nM) of KR127 was obtained by mutational analysis of the heavy chain CDR3 (HCDR3) of KR127. The AP2 antibody was 4-fold more sensitive in detecting the S1-tagged protein than KR127. The S1 tag-KR127 or AP2 combination could be universally used for monitoring protein expression, localizing proteins, and protein purification, as well as studying protein interactions.-
dc.publisherElsevier-
dc.titleA new epitope tag from hepatitis B virus preS1 for immunodetection, localization and affinity purification of recombinant proteins-
dc.title.alternativeA new epitope tag from hepatitis B virus preS1 for immunodetection, localization and affinity purification of recombinant proteins-
dc.typeArticle-
dc.citation.titleJournal of Immunological Methods-
dc.citation.number1-
dc.citation.endPage89-
dc.citation.startPage77-
dc.citation.volume283-
dc.contributor.affiliatedAuthorC Y Maeng-
dc.contributor.affiliatedAuthorS H Min-
dc.contributor.affiliatedAuthorM H Jang-
dc.contributor.affiliatedAuthorS O Yoon-
dc.contributor.affiliatedAuthorJ H Kim-
dc.contributor.affiliatedAuthorHyo Jeong Hong-
dc.contributor.alternativeName오미숙-
dc.contributor.alternativeName김근수-
dc.contributor.alternativeName장영국-
dc.contributor.alternativeName맹철영-
dc.contributor.alternativeName민순홍-
dc.contributor.alternativeName장명희-
dc.contributor.alternativeName윤선옥-
dc.contributor.alternativeName김정희-
dc.contributor.alternativeName홍효정-
dc.identifier.bibliographicCitationJournal of Immunological Methods, vol. 283, no. 1, pp. 77-89-
dc.identifier.doi10.1016/j.jim.2003.08.010-
dc.subject.keywordAffinity maturation-
dc.subject.keywordHepatitis B virus preS1-
dc.subject.keywordMonoclonal antibody-
dc.subject.keywordProtein expression-
dc.subject.keywordProtein tagging-
dc.subject.keywordProteomics-
dc.subject.localaffinity maturation-
dc.subject.localaffinity Maturation-
dc.subject.localAffinity maturation-
dc.subject.localhepatitis B virus preS1-
dc.subject.localHepatitis B virus preS1-
dc.subject.localMonoclonal antibody (mAb)-
dc.subject.localMonoclonal antibodies-
dc.subject.localMonoclonal Antibodies-
dc.subject.localMonoclonal antibody-
dc.subject.localmonoclonal antibody-
dc.subject.localMonoclonal Antibody-
dc.subject.localmonoclonal antibodies-
dc.subject.localProtein expression-
dc.subject.localprotein expression-
dc.subject.localProtein tagging-
dc.subject.localProteomic-
dc.subject.localProteomics-
dc.description.journalClassY-
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