Mutations in PMR1 stimulate xylose isomerase activity and anaerobic growth on xylose of engineered Saccharomyces cerevisiae by influencing manganese homeostasis

Maarten D Verhoeven, Misun Lee, Lycka Kamoen, Marcel van den Broek, Dick B Janssen, Jean-Marc G Daran, Antonius J A van Maris, Jack T Pronk

OnderzoeksoutputAcademicpeer review

33 Citaten (Scopus)
244 Downloads (Pure)

Samenvatting

Combined overexpression of xylulokinase, pentose-phosphate-pathway enzymes and a heterologous xylose isomerase (XI) is required but insufficient for anaerobic growth of Saccharomyces cerevisiae on d-xylose. Single-step Cas9-assisted implementation of these modifications yielded a yeast strain expressing Piromyces XI that showed fast aerobic growth on d-xylose. However, anaerobic growth required a 12-day adaptation period. Xylose-adapted cultures carried mutations in PMR1, encoding a Golgi Ca(2+)/Mn(2+) ATPase. Deleting PMR1 in the parental XI-expressing strain enabled instantaneous anaerobic growth on d-xylose. In pmr1 strains, intracellular Mn(2+) concentrations were much higher than in the parental strain. XI activity assays in cell extracts and reconstitution experiments with purified XI apoenzyme showed superior enzyme kinetics with Mn(2+) relative to other divalent metal ions. This study indicates engineering of metal homeostasis as a relevant approach for optimization of metabolic pathways involving metal-dependent enzymes. Specifically, it identifies metal interactions of heterologous XIs as an underexplored aspect of engineering xylose metabolism in yeast.
Originele taal-2English
Artikelnummer46155
Aantal pagina's11
TijdschriftScientific Reports
Volume7
DOI's
StatusPublished - 12-apr-2017

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