Optimization of Bacillus α‐amylase production by Saccharomyces cerevisiae

L. Ruohonen, M. Penttilä, S. Keränen

Research output: Contribution to journalArticleScientificpeer-review

26 Citations (Scopus)

Abstract

Production of Bacillus amyloliquefaciens α‐amylase by Saccharomyces cerevisiae using the multicopy plasmid pAAH5 and ways of improving the yields of secreted enzyme were studied. In standard non‐buffered medium, α‐amylase was rapidly inactivated but stabilization of the pH at 6 led to stable accumulation of α‐amylase in the culture medium. Removal of 1100 bp of the upstream sequence of the ADH1 promoter present on pAAH5 resulted in delayed but increased α‐amylase production: 29‐fold in selective medium, two‐fold in non‐selective medium. With the original ADH1 promoter, accumulation of α‐amylase in the medium started to level off before the cultures reached stationary phase and was very low when exponentially growing cells were transferred from glucose to ethanol. This coincided with the appearance of a mRNA larger than the α‐amylase messenger. With the shortened promoter, the normal‐size α‐amylase mRNA was detected under all growth conditions and α‐amylase was efficiently secreted into the medium also late in stationary phase and after transfer to ethanol. Highest total yields of α‐amylase were obtained with the short promoter in non‐selective glucose‐containing medium; this may be explained by the greater final cell density obtained. However, the production of α‐amylase per cell mass was higher in ethanol‐containing selective medium. Seventy to eighty per cent of the α‐amylase activity was secreted into the medium independent of the total amount produced.

Original languageEnglish
Pages (from-to)337-346
Number of pages10
JournalYeast
Volume7
Issue number4
DOIs
Publication statusPublished - 1 Jan 1991
MoE publication typeA1 Journal article-refereed

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Amylases
Bacilli
Yeast
Bacillus
Saccharomyces cerevisiae
Ethanol
Messenger RNA
Glucose
Culture Media
Plasmids
Enzymes
Stabilization
Cell Count

Keywords

  • ADHI promoter
  • heterologous expression
  • increased production
  • regulation
  • Yeast

Cite this

Ruohonen, L. ; Penttilä, M. ; Keränen, S. / Optimization of Bacillus α‐amylase production by Saccharomyces cerevisiae. In: Yeast. 1991 ; Vol. 7, No. 4. pp. 337-346.
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Optimization of Bacillus α‐amylase production by Saccharomyces cerevisiae. / Ruohonen, L.; Penttilä, M.; Keränen, S.

In: Yeast, Vol. 7, No. 4, 01.01.1991, p. 337-346.

Research output: Contribution to journalArticleScientificpeer-review

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