Increasing Heterologous Protein Secretion From the Yeast S. Cerevisiae Through Manipulation of Cellular Redox Factors
Bannister, Sarah Jane
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https://hdl.handle.net/2142/82479
Description
Title
Increasing Heterologous Protein Secretion From the Yeast S. Cerevisiae Through Manipulation of Cellular Redox Factors
Author(s)
Bannister, Sarah Jane
Issue Date
2000
Doctoral Committee Chair(s)
Wittrup, K. Dane
Department of Study
Chemical Engineering
Discipline
Chemical Engineering
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Biology, Microbiology
Language
eng
Abstract
The work described in this thesis involves the effect of cellular redox factors on BPTI secretion. Glutathione levels and redox state were altered in an attempt to determine if optimal BPTI secretion requires a certain total glutathione level and/or redox state. Evidence for an optimal glutathione level that results in high BPTI secretion was obtained. Thioredoxin deficiency was also shown to increase BPTI secretion. Several possible actors in the ER oxidative protein folding chain were identified and their effects on BPTI secretion measured. Overexpression of Ero1p was shown to increase BPTI secretion, while overexpression of flavin-containing monooxygenase (FMO) and Fun9p did not. Overexpression of two potential sulfhydryl oxidases, Erv1p and Erv2p, did not increase BPTI secretion. The addition of ubiquinone, which has been shown to be an electron acceptor from the E. coli disulfide bond formation chain, increased BPTI secretion from cultures overexpressing FMO, but under no other scenarios. A system has been set up to utilize the yeast surface display system to screen a cDNA library for factors that increase BPTI surface display, and therefore secretion. The information gathered in this work identifies new directions for increasing protein secretion in yeast.
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