Enzymic analysis of NADPH metabolism in β-lactam-producing Penicillium chrysogenum: Presence of a mitochondrial NADPH dehydrogenase

Diana M. Harris, Jasper A. Diderich, Zita A. Van Der Krogt, Marijke A H Luttik, Léonie M. Raamsdonk, Roel A L Bovenberg, Walter M. Van Gulik, Johannes P. Van Dijken, Jack T. Pronk

Research output: Contribution to journalArticleScientificpeer-review

Abstract

Based on assumed reaction network structures, NADPH availability has been proposed to be a key constraint in β-lactam production by Penicillium chrysogenum. In this study, NADPH metabolism was investigated in glucose-limited chemostat cultures of an industrial P. chrysogenum strain. Enzyme assays confirmed the NADP+-specificity of the dehydrogenases of the pentose-phosphate pathway and the presence of NADP+-dependent isocitrate dehydrogenase. Pyruvate decarboxylase/NADP+-linked acetaldehyde dehydrogenase and NADP+-linked glyceraldehyde-3- phosphate dehydrogenase were not detected. Although the NADPH requirement of penicillin-G-producing chemostat cultures was calculated to be 1.4-1.6-fold higher than that of non-producing cultures, in vitro measured activities of the major NADPH-providing enzymes were the same. Isolated mitochondria showed high rates of antimycin A-sensitive respiration of NADPH, thus indicating the presence of a mitochondrial NADPH dehydrogenase that oxidises cytosolic NADPH. The presence of this enzyme in P. chrysogenum might have important implications for stoichiometric modelling of central carbon metabolism and β-lactam production and may provide an interesting target for metabolic engineering.

Original languageEnglish
Pages (from-to)91-101
JournalMetabolic Engineering
Volume8
Issue number2
DOIs
Publication statusPublished - 2006

Keywords

  • Chemostat cultures
  • Metabolic engineering
  • Mitochondria
  • NADPH
  • Penicillin-G
  • Penicillium chrysogenum
  • Redox metabolism
  • Respiratory chain

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