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Title:The role of BIP and PDI in the secretion of foreign proteins in the yeast Saccharomyces cerevisiae
Author(s):Robinson, Anne Skaja
Doctoral Committee Chair(s):Lauffenburger, Douglas A.
Department / Program:Chemical and Biomolecular Engineering
Discipline:Chemical and Biomolecular Engineering
Degree Granting Institution:University of Illinois at Urbana-Champaign
Subject(s):Biology, Cell
Chemistry, Biochemistry
Engineering, Chemical
Abstract:As a single-celled microbial eucaryotic host for protein expression, the yeast Saccharomyces cerevisiae offers some of the advantages of bacterial systems, such as ease of fermentation, and of eucaryotic systems, such as post-translational modifications of secreted proteins. One disadvantage, however, is that secretion of foreign proteins is generally inefficient.
The limiting step in protein secretion is often protein folding in the lumen of the endoplasmic reticulum (ER), a process assisted by accessory factors resident in this compartment. Chaperones, such as the hsp70 homolog binding protein (BiP), bind reversibly to the unfolded conformation of proteins, preventing irreversible aggregation. Foldases, such as protein disulfide isomerase (PDI), catalyze the formation and rearrangement of bonds which stabilize the folded conformation of proteins. I have examined the role of BiP and PDI in determining the efficiency of foreign protein secretion in the yeast Saccharomyces cerevisiae.
I have found that high-level expression of foreign genes does not always lead to increased production of foreign proteins. In fact, prolonged constitutive expression of foreign secreted proteins reduces soluble BiP and PDI protein to levels undetectable by Western immunoassay. Fifteen-fold overexpression of PDI from a strong glycolytic promoter results in significant enhancement of secretion for some foreign proteins. Improved secretion is correlated with decreased ER retention, indicative of accelerated folding. When the chromosomal copy of BiP is deleted, and BiP levels are tightly regulated from a plasmid-borne copy of the gene controlled by the CUP1 promoter, both secretion and growth are decreased significantly when BiP falls below wild type levels. A mechanistic model can account for the behavior of BiP experimentally, and predictions have been made for altering cellular properties to increase protein secretion.
Issue Date:1994
Rights Information:Copyright 1994 Robinson, Anne Skaja
Date Available in IDEALS:2011-05-07
Identifier in Online Catalog:AAI9512530
OCLC Identifier:(UMI)AAI9512530

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