Engineering heterologous molybdenum-cofactor-biosynthesis and nitrate-assimilation pathways enables nitrate utilization by Saccharomyces cerevisiae

A Multiphase Multiobjective Dynamic Genome-Scale Model Shows Different Redox Balancing among Yeast Species of the Saccharomyces Genus in Fermentation

“…Yeasts have been used to produce fermented foods and beverages for millennia and are among the most frequently used microorganisms in biotechnology. Saccharomyces cerevisiae dominates the scene and many research efforts focus on engineering this species for particular applications (for examples, see references 1 to 4 ). Nowadays it is used to produce glycerol ( 5 ), biopharmaceutical proteins ( 6 ), or secondary metabolites, such as aromas or bioflavors ( 7 , 8 ).…”

Engineering of molybdenum-cofactor-dependent nitrate assimilation in Yarrowia lipolytica

“…Based on a previous work in S. cerevisiae (Perli et al . 2021 ), it is not fewer than 11 genes from the yeast O. parapolymorpha and the algae C. reinhardtii that would be required to introduce Moco biosynthesis and nitrate assimilation in Y. lipolytica . The gene-set comprises seven genes coding for Moco biosynthesis proteins ( OpCNX1 , OpCNX2 , OpCNX3 , OpCNX4 , OpCNX5 , OpCNX6 and OpNFS1 ), three genes coding for the nitrate assimilation pathway ( OpYNT1 , OpYNR1 and OpYNI1 ) from O. parapolymorpha , and one gene coding for the high-affinity molybdate transporter ( CrMoT1 ) from C. reinhardtii (Fig.…”

“… 2013 ; Perli et al . 2021 ) BioProject PRJNA60503). A gene coding for a previously characterized high-affinity molybdenum transporter, CrMoT1 , from C. reinhardtii was also included in the gene-set (Tejada-Jimenez et al .…”

“…Cell extract preparation and activity measurements were performed as previously described (Perli et al . 2021 ). In brief, frozen stock cultures of each tested strain and evolved populations were used to inoculate 20 mL starter cultures, which were then used to inoculate 100 mL shake flask cultures on the same medium, to an initial OD 660 of 0.2.…”

“…More recently, the molybdenum cofactor biosynthesis pathway from the methylotrophic yeast Ogataea parapolymorpha was introduced in S. cerevisiae allowing expression of a functional Moco-dependent nitrate reductase that could support growth on media containing nitrate as sole nitrogen source (Perli et al . 2021 ). Although these pioneering studies demonstrated how metabolic landscape could be expanded beyond the natural ability of a microorganism, these approaches have not yet been transposed in other industrially relevant yeast species.…”