RT Journal Article
SR Electronic
T1 Drop-In Biofuel production by using fatty acid photodecarboxylase from <em>Chlorella variabilis</em> in the oleaginous yeast <em>Yarrowia lipolytica</em>
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 468876
DO 10.1101/468876
A1 Stefan Bruder
A1 Eva Johanna Moldenhauer
A1 Robert Denis Lemke
A1 Rodrigo Ledesma-Amaro
A1 Johannes Kabisch
YR 2019
UL http://biorxiv.org/content/early/2019/04/27/468876.abstract
AB Background Oleaginous yeasts are potent hosts for the renewable production of lipids and harbor great potential for derived products, such as biofuels. Several promising processes have been described that produce hydrocarbon drop-in biofuels based on fatty acid decarboxylation and fatty aldehyde decarbonylation. Unfortunately, besides fatty aldehyde toxicity and high reactivity, the most investigated enzyme, aldehyde-deformylating oxygenase, shows unfavorable catalytic properties which hindered high yields in previous metabolic engineering approaches.Results To demonstrate an alternative alkane production pathway for oleaginous yeasts, we describe the production of diesel-like, odd-chain alkanes and alkenes, by heterologously expressing a recently discovered light-driven oxidase from Chlorella variabilis (CvFAP) in Yarrowia lipolytica. Initial experiments showed that only strains engineered to have an increased pool of free fatty acids showed to be susceptible to sufficient decarboxylation. Providing these strains with glucose and light in a synthetic medium resulted in titers of 10.9 mg/L of hydrocarbons. Using custom 3D printed labware for lighting bioreactors, and an automated pulsed glycerol fed-batch strategy, intracellular titers of 58.7 mg/L were achieved.Conclusions Oleaginous yeasts such as Yarrowia lipolytica can transform renewable resources such as glycerol into fatty acids and lipids. By heterologously expressing a fatty acid photodecarboxylase from the algae Chlorella variabilis hydrocarbons were produced in several scales from microwell plate to 400 ml bioreactors. The developed bioprocess shows a route to the renewable production of hydrocarbons for a variety of applications ranging from representing a substrate for further enzymatic or chemical modification or as a drop-in biofuel blend.Short abstract Oleaginous yeasts are potent hosts for the renewable production of lipids, fatty acids and derived products such as biofuels. Here, we describe, the production of odd-numbered alkanes and alkenes with a length of 17 and 15 carbons by expression of a fatty acid photodecarboxylase (CvFAP) from Chlorella variabilis in different Yarrowia lipolytica strains under different regimes of blue light exposure in several scales from microwell plate to 400 ml bioreactors.List of abbreviationsÅÅngströmAAamino acidAAR (FAR)(fatty) acyl-ACP reductaseADOaldehyde-deformylating oxygenaseCARcarboxylic acid reductaseCdwcell dry weightCv/rChlorella variabilis/reinhardtiiDOdissolved oxygenDOXα-dioxygenaseFFAfree fatty acidsGMCglucose-methanol-cholineSCOsingle cell oilFAPfatty acid photodecarboxylaseFFAfree fatty acidsRTroom temperatureYaliTARtransformation-associated recombination assisted by Y. lipolyticaWTwild type