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2016 Spring- Tzu-Chiang Han Dissertation (Updated II) .pdf (4.49 MB)
ETD Abstract Container
Abstract Header
Control of Intein-Mediated Self-Cleaving Tag for Recombinant Protein Purification
Author Info
Han, Tzu-Chiang
ORCID® Identifier
http://orcid.org/0000-0001-9942-1646
Permalink:
http://rave.ohiolink.edu/etdc/view?acc_num=osu1453206997
Abstract Details
Year and Degree
2016, Doctor of Philosophy, Ohio State University, Chemical Engineering.
Abstract
Recent advances in upstream bioprocessing have largely increased the productivities of recombinant protein expression in various expression systems. This has led to an increasingly urgent need for rapid, cost-effective and large-scale downstream unit operations. Affinity tag-based purification provides an efficient platform to purify virtually any uncharacterized protein in a highly selective, high-throughput manner. With the combination of a self-cleaving intein, the purification and the tag removal processes can all be carried out in a single chromatography step, thus dramatically reducing the cost of downstream processing. Over the past decades, inteins have been genetically modified for better controllability and cleaving ability. However, unwanted in vivo premature intein cleaving remains a major bottleneck for applications in mammalian systems. In this dissertation, recent progresses in the control of intein cleaving will be discussed. A continuous delta I-CM intein and a zinc binding motif fusion allow the intein-tagged precursor proteins to be expressed in mammalian systems with less than 40-50% in vivo premature cleaving in the presence of 200 micro molar zinc chloride. An alternative, the Npu split intein system, has been shown to completely eliminate premature cleaving and has been altered into a pH sensitive cleaving intein. These efforts to control intein cleaving have led to the development of a robust and efficient platform technology for non-antibody recombinant protein downstream purification.
Committee
David Wood (Advisor)
Andre Palmer (Committee Member)
Shang-Tian Yang (Committee Member)
Werner Tajrks (Committee Member)
Pages
179 p.
Subject Headings
Chemical Engineering
Keywords
recombinant protein purification
;
intein
;
mammalian cell
;
tag removal
;
downstream process
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Citations
Han, T.-C. (2016).
Control of Intein-Mediated Self-Cleaving Tag for Recombinant Protein Purification
[Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1453206997
APA Style (7th edition)
Han, Tzu-Chiang.
Control of Intein-Mediated Self-Cleaving Tag for Recombinant Protein Purification.
2016. Ohio State University, Doctoral dissertation.
OhioLINK Electronic Theses and Dissertations Center
, http://rave.ohiolink.edu/etdc/view?acc_num=osu1453206997.
MLA Style (8th edition)
Han, Tzu-Chiang. "Control of Intein-Mediated Self-Cleaving Tag for Recombinant Protein Purification." Doctoral dissertation, Ohio State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=osu1453206997
Chicago Manual of Style (17th edition)
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Document number:
osu1453206997
Download Count:
2,278
Copyright Info
© 2016, all rights reserved.
This open access ETD is published by The Ohio State University and OhioLINK.