cff-version: 1.2.0
abstract: "<p>Metabolic capabilities of cells are not only defined by their
repertoire of enzymes and metabolites, but also by availability of enzyme
cofactors. The molybdenum cofactor (Moco) is widespread among eukaryotes but
absent from the industrial yeast <i>Saccharomyces</i> <i>cerevisiae</i>. In this study, we identified 7 Moco biosynthesis genes
in the non-conventional yeast <i>Ogataea parapolymorpha</i> by <i>Spy</i>cas9-mediated
mutational analysis and expressed them in <i>S.
cerevisiae</i>. Functionality of the heterologously expressed<i> </i>Moco biosynthesis pathway in <i>S. cerevisiae</i> was assessed by
co-expressing <i>O. parapolymorpha </i>nitrate-assimilation
enzymes, including the Moco-dependent nitrate reductase. Following two-weeks of
incubation, growth of the engineered <i>S.
cerevisiae</i> was observed on nitrate as sole nitrogen source. Relative to the
engineered, evolved nitrate-assimilating <i>S.
cerevisiae</i> strains isolated from these cultures showed increased copy
numbers of the heterologous genes, increased levels of the encoded proteins and
a 5-fold higher nitrate-reductase activity in cell extracts. Growth at nM molybdate
concentrations was enabled by co-expression of a <i>Chlamydomonas reinhardtii</i> high-affinity molybdate transporter. In
serial batch cultures on nitrate-containing medium, a non-engineered <i>S. cerevisiae</i> was rapidly outcompeted by
the spoilage yeast <i>Brettanomyces bruxellensis. </i>In contrast, an engineered and evolved nitrate-assimilating <i>S.
cerevisiae</i> strains persisted during 35 generations of co-cultivation.
This result indicates that the ability of engineered strains to use nitrate may
be applicable to improve competitiveness industrial processes upon
contamination with spoilage yeasts. Since over 50 Moco-dependent enzymes have
been described, introduction of a functional Moco synthesis pathway offers interesting
options to further broaden the biocatalytic repertoire of <i>S. cerevisiae</i>. </p>"
authors:
  - family-names: Daran
    given-names: Jean-Marc
    orcid: "https://orcid.org/0000-0003-3136-8193"
  - family-names: Perli
    given-names: T. (Thomas)
    orcid: "https://orcid.org/0000-0002-4910-3292"
  - family-names: Vorm  van der
    given-names: Daan N. A.
  - family-names: Wassink
    given-names: Mats
  - family-names: van den Broek
    given-names: Marcel
  - family-names: Pronk
    given-names: Jack T.
title: "Data underlying research on expression of heterologous molybdenum-cofactor-biosynthesis and nitrate-assimilation genes enables nitrate utilization by Saccharomyces cerevisiae"
keywords:
version: 1
identifiers:
  - type: doi
    value: 10.4121/13194518.v1
license: CC BY-NC-ND 4.0
date-released: 2020-12-02