| Engineering and refolding of a novel trimeric fusion protein TRAIL-collagen XVIII NC1. | |
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MedLine Citation:
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PMID: 23208613 Owner: NLM Status: Publisher |
Abstract/OtherAbstract:
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Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is considered to be a promising anticancer agent because its active form TRAIL trimer is able to induce apoptosis in different tumor cell lines while sparing normal cells. However, TRAIL trimer possesses a short half-life and low stability, which turns out to be a major obstacle for the development of clinical trials. In our present study, we constructed a recombined TRAIL trimer by genetic fusion of non-collagenous domain (NC1) of human collagen XVIII or its trimerization domain (TD) to C-terminus of TRAIL via a flexible linker, and then refolded the fusion proteins using a two-step refolding approach, namely a combination of dilution and gel filtration chromatography. As a result, both recombinant proteins, TRAIL-NC1 and TRAIL-TD, were expressed in Escherichia coli as inclusion bodies, and they exhibited difficultly to refold efficiently by conventional methods. Thereby, we applied a modified two-step refolding approach to refold fusion proteins. More than 55 % of TRAIL-NC1 and 90 % of TRAIL-TD protein activity was recovered during the two-step refolding approach, and their stability was also increased significantly. Also, size exclusion chromatography showed refolded TRAIL-NC1 was a trimer while TRAIL-TD, hexamer. However, both of them exerted good apoptosis activity on NCI-H460 cells. |
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Authors:
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Li Qiang Pan; Zhang Ming Xie; Xiao Jing Tang; Min Wu; Fu Rong Wang; Hua Naranmandura; Shu Qing Chen |
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Publication Detail:
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Type: JOURNAL ARTICLE Date: 2012-12-4 |
Journal Detail:
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Title: Applied microbiology and biotechnology Volume: - ISSN: 1432-0614 ISO Abbreviation: Appl. Microbiol. Biotechnol. Publication Date: 2012 Dec |
Date Detail:
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Created Date: 2012-12-4 Completed Date: - Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 8406612 Medline TA: Appl Microbiol Biotechnol Country: - |
Other Details:
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Languages: ENG Pagination: - Citation Subset: - |
Affiliation:
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Department of Pharmacology, Toxicology and Biochemical Pharmaceutics, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou, 310058, China. |
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From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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