Two-step chromatographic purification of glutathione S-transferase-tagged human papillomavirus type 16 E6 protein and its application for serology
DC Field | Value | Language |
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dc.contributor.author | Xu, Mei Ling | - |
dc.contributor.author | Kim, Seung Cheol | - |
dc.contributor.author | Kim, Hyoung Jin | - |
dc.contributor.author | Ju, Woong | - |
dc.contributor.author | Kim, Yun Hwan | - |
dc.contributor.author | Kim, Hong-Jin | - |
dc.date.available | 2019-03-08T08:58:12Z | - |
dc.date.issued | 2017-04 | - |
dc.identifier.issn | 1046-5928 | - |
dc.identifier.issn | 1096-0279 | - |
dc.identifier.uri | https://scholarworks.bwise.kr/cau/handle/2019.sw.cau/4639 | - |
dc.description.abstract | Human papillomavirus (HPV) E6 protein is an oncoprotein with a pivotal role in cervical carcinogenesis. Expression and purification of HPV E6 from Escherichia coli (E. coli) has been difficult because of its strong hydrophobicity even when expressed as a fusion protein with glutathione S-transferase (GST). There has been no protocol suggested for purifying GST-tagged HPV E6 protein with high purity so far. Herein, we provide efficient protocol for purifying GST-HPV16 E6 protein for the first time. In the current study, the GST-tagged protein was expressed in E. coli and a purification method was designed using cation exchange chromatography followed by GST-affinity chromatography. Using physiological pH buffer during cell lysis and first cation-exchange chromatography significantly reduced yield of full-length GST-HPV16 E6 protein. It was found that using an alkaline buffer during cation-exchange chromatography was needed to obtain full length GST-HPV16 E6 protein. GST-HPV16 E6 protein recovered from the purification using alkaline condition retained its inherent p53-binding ability. Moreover, we were able to detect anti-HPV16 E6 antibodies with high sensitivity in sera from patients with cervical cancer using the GST HPV16 E6 protein. It was found that the GST-HPV16 E6 protein could be used as a coating agent to enhance the sensitivity of detection of serum anti-HPV16 E6 antibodies when treated with ethylene glycol-bis (beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA). These results indicate that the two-step chromatographic purification allows obtaining high purity of GST-HPV16 E6 protein and the GST-HPV16 E6 is suitable to be used as an antigen of serology assay. (C) 2017 Elsevier Inc. All rights reserved. | - |
dc.format.extent | 8 | - |
dc.language | 영어 | - |
dc.language.iso | ENG | - |
dc.publisher | ACADEMIC PRESS INC ELSEVIER SCIENCE | - |
dc.title | Two-step chromatographic purification of glutathione S-transferase-tagged human papillomavirus type 16 E6 protein and its application for serology | - |
dc.type | Article | - |
dc.identifier.doi | 10.1016/j.pep.2017.01.004 | - |
dc.identifier.bibliographicCitation | PROTEIN EXPRESSION AND PURIFICATION, v.132, pp 19 - 26 | - |
dc.description.isOpenAccess | N | - |
dc.identifier.wosid | 000401298600003 | - |
dc.identifier.scopusid | 2-s2.0-85009223601 | - |
dc.citation.endPage | 26 | - |
dc.citation.startPage | 19 | - |
dc.citation.title | PROTEIN EXPRESSION AND PURIFICATION | - |
dc.citation.volume | 132 | - |
dc.type.docType | Article | - |
dc.publisher.location | 미국 | - |
dc.subject.keywordAuthor | Human papillomavirus | - |
dc.subject.keywordAuthor | E6 protein | - |
dc.subject.keywordAuthor | Serology | - |
dc.subject.keywordAuthor | Cervical cancer | - |
dc.subject.keywordAuthor | Glutathione S-transferase | - |
dc.subject.keywordPlus | CERVICAL INTRAEPITHELIAL NEOPLASIA | - |
dc.subject.keywordPlus | ESCHERICHIA-COLI | - |
dc.subject.keywordPlus | CANCER | - |
dc.subject.keywordPlus | ONCOPROTEIN | - |
dc.subject.keywordPlus | TRANSFORMATION | - |
dc.subject.keywordPlus | KERATINOCYTES | - |
dc.subject.keywordPlus | DEGRADATION | - |
dc.subject.keywordPlus | ANTIBODIES | - |
dc.subject.keywordPlus | ABILITY | - |
dc.subject.keywordPlus | LESIONS | - |
dc.relation.journalResearchArea | Biochemistry & Molecular Biology | - |
dc.relation.journalResearchArea | Biotechnology & Applied Microbiology | - |
dc.relation.journalWebOfScienceCategory | Biochemical Research Methods | - |
dc.relation.journalWebOfScienceCategory | Biochemistry & Molecular Biology | - |
dc.relation.journalWebOfScienceCategory | Biotechnology & Applied Microbiology | - |
dc.description.journalRegisteredClass | sci | - |
dc.description.journalRegisteredClass | scie | - |
dc.description.journalRegisteredClass | scopus | - |
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