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Comparison of three expression systems for heterologous xylanase production by S-cerevisiae in defined medium

Görgens, Johann F; Pianas, Jordi; van Zyl, W H; Knoetze, J H and Hahn-Hägerdal, Bärbel LU (2004) In Yeast 21(14). p.1205-1217
Abstract
The influence of the auxotrophic deficiencies of the host strain and expression vector selection on the production of a heterologous protein was investigated. Heterologous xylanase production by two prototrophic S. cerevisiae transformants, containing either a plasmid-based, YEp-type expression system or an integrative, YIp-type expression system, were compared with production by an auxotrophic transformant, containing an identical YEp-type expression system, in batch and continuous cultivation, using a chemically defined medium. Heterologous xylanase production by the auxotrophic strains in defined medium was critically dependent on the availability of amino acids, as extracellular xylanase production increased dramatically when amino... (More)
The influence of the auxotrophic deficiencies of the host strain and expression vector selection on the production of a heterologous protein was investigated. Heterologous xylanase production by two prototrophic S. cerevisiae transformants, containing either a plasmid-based, YEp-type expression system or an integrative, YIp-type expression system, were compared with production by an auxotrophic transformant, containing an identical YEp-type expression system, in batch and continuous cultivation, using a chemically defined medium. Heterologous xylanase production by the auxotrophic strains in defined medium was critically dependent on the availability of amino acids, as extracellular xylanase production increased dramatically when amino acids were over-consumed from the medium to the point of saturating the cell. Saturation with amino acids, indicated by an increased leakage of amino acids from the cell, was thus a prerequisite for high level of heterologous protein production by the auxotrophic strain. Maximal xylanase production levels by the auxotrophic strain corresponded to the levels obtained with a similar prototrophic strain during cultivation in defined medium without amino acids. Superfluous auxotrophic markers thus had a strong deleterious effect on heterologous protein production by recombinant yeasts, and the use of such strains should be limited to initial exploratory investigations. The increased copy number and foreign gene dosage of the YEp-based expression vector, stabilized by the ura3 furI autoselection system, significantly improved production levels of heterollogous xylanase, compared to the YIp system, which is based on a single integration into the yeast genome. No evidence was found of the possible saturation of the host secretory capacity by multicopy overexpression. Stable production of heterologous xylanase at high levels by the prototrophic YEp-based recombinant strain, compared to the YIp system, was demonstrated. Copyright (C) 2004 John Wiley Sons, Ltd. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Yeast
volume
21
issue
14
pages
1205 - 1217
publisher
John Wiley & Sons
external identifiers
  • pmid:15515128
  • wos:000225236100003
  • scopus:9144226306
ISSN
1097-0061
DOI
10.1002/yea.1175
language
English
LU publication?
yes
id
51a4becd-278d-4a0e-bafa-2679e29db3f2 (old id 140456)
date added to LUP
2007-06-28 11:43:48
date last changed
2017-03-19 04:15:05
@article{51a4becd-278d-4a0e-bafa-2679e29db3f2,
  abstract     = {The influence of the auxotrophic deficiencies of the host strain and expression vector selection on the production of a heterologous protein was investigated. Heterologous xylanase production by two prototrophic S. cerevisiae transformants, containing either a plasmid-based, YEp-type expression system or an integrative, YIp-type expression system, were compared with production by an auxotrophic transformant, containing an identical YEp-type expression system, in batch and continuous cultivation, using a chemically defined medium. Heterologous xylanase production by the auxotrophic strains in defined medium was critically dependent on the availability of amino acids, as extracellular xylanase production increased dramatically when amino acids were over-consumed from the medium to the point of saturating the cell. Saturation with amino acids, indicated by an increased leakage of amino acids from the cell, was thus a prerequisite for high level of heterologous protein production by the auxotrophic strain. Maximal xylanase production levels by the auxotrophic strain corresponded to the levels obtained with a similar prototrophic strain during cultivation in defined medium without amino acids. Superfluous auxotrophic markers thus had a strong deleterious effect on heterologous protein production by recombinant yeasts, and the use of such strains should be limited to initial exploratory investigations. The increased copy number and foreign gene dosage of the YEp-based expression vector, stabilized by the ura3 furI autoselection system, significantly improved production levels of heterollogous xylanase, compared to the YIp system, which is based on a single integration into the yeast genome. No evidence was found of the possible saturation of the host secretory capacity by multicopy overexpression. Stable production of heterologous xylanase at high levels by the prototrophic YEp-based recombinant strain, compared to the YIp system, was demonstrated. Copyright (C) 2004 John Wiley Sons, Ltd.},
  author       = {Görgens, Johann F and Pianas, Jordi and van Zyl, W H and Knoetze, J H and Hahn-Hägerdal, Bärbel},
  issn         = {1097-0061},
  language     = {eng},
  number       = {14},
  pages        = {1205--1217},
  publisher    = {John Wiley & Sons},
  series       = {Yeast},
  title        = {Comparison of three expression systems for heterologous xylanase production by S-cerevisiae in defined medium},
  url          = {http://dx.doi.org/10.1002/yea.1175},
  volume       = {21},
  year         = {2004},
}