Mitochondria from Ogataea parapolymorpha harbor a branched electron-transport chain containing a proton-pumping Complex I NADH dehydrogenase and three Type II NADH dehydrogenases (NDH-2). To investigate the physiological role, localization and substrate specificity of these enzymes, the growth of various NADH dehydrogenase knockout mutants was quantitatively characterized in shake-flask and chemostat cultures, followed by oxygen-uptake experiments with isolated mitochondria. NAD(P)H:quinone oxidoreduction of the three NDH-2 were individually assessed. Our findings reveal that the O. parapolymorpha respiratory chain contains an internal NADH-accepting NDH-2 (Ndh2-1/OpNdi1), at least one external NAD(P)H-accepting enzyme, and likely additional mechanisms for respiration-linked oxidation of cytosolic NADH. Metabolic regulation appears to prevent competition between OpNdi1 and Complex I for mitochondrial NADH. With the exception of OpNdi1, the respiratory chain of O. parapolymorpha exhibits metabolic redundancy and tolerates deletion of multiple NADH-dehydrogenase genes without compromising fully respiratory metabolism.@en

A novel fermentation process was developed in which renewable electricity is indirectly used as an energy source in fermentation, synergistically decreasing both the consumption of sugar as a first generation carbon source and emission of the greenhouse gas CO₂. As an illustration, a glucose-based process is co-fed with formic acid, which can be generated by capturing CO₂ from fermentation offgas followed by electrochemical reduction with renewable electricity. This “closed carbon loop” concept is demonstrated by a case study in which cofeeding formic acid is shown to significantly increase the yield of biomass on glucose of the industrially relevant yeast species Yarrowia lipolytica. First, the optimal feed ratio of formic acid to glucose is established using chemostat cultivations. Subsequently, guided by a dynamic fermentation process model, a fed-batch protocol is developed and demonstrated on laboratory scale. Finally, the developed fed-batch process is tested and proven to be scalable at pilot scale. Extensions of the concept are discussed to apply the concept to anaerobic fermentations, and to recycle the O₂ that is co-generated with the formic acid to aerobic fermentation processes for intensification purposes.

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