Engineering of a high lipid producing Yarrowia lipolytica strain.

Emily H Greenhagen,Gregory Stephanopoulos,Jonathan Friedlander,Rebecca L Nugent,Colin R South,Jonathan Afshar,Kyle Macewen,Maureen A Hamilton,Annapurna Kamineni,A Joe Shaw,Vasiliki Tsakraklides,Andrew L Consiglio,Elena E Brevnova,Donald V Crabtree

doi:10.1186/s13068-016-0492-3

Abstract

BackgroundMicrobial lipids are produced by many oleaginous organisms including the well-characterized yeast Yarrowia lipolytica, which can be engineered for increased lipid yield by up-regulation of the lipid biosynthetic pathway and down-regulation or deletion of competing pathways.ResultsWe describe a strain engineering strategy centered on diacylglycerol acyltransferase (DGA) gene overexpression that applied combinatorial screening of overexpression and deletion genetic targets to construct a high lipid producing yeast biocatalyst. The resulting strain, NS432, combines overexpression of a heterologous DGA1 enzyme from Rhodosporidium toruloides, a heterlogous DGA2 enzyme from Claviceps purpurea, and deletion of the native TGL3 lipase regulator. These three genetic modifications, selected for their effect on lipid production, enabled a 77 % lipid content and 0.21 g lipid per g glucose yield in batch fermentation. In fed-batch glucose fermentation NS432 produced 85 g/L lipid at a productivity of 0.73 g/L/h.ConclusionsThe yields, productivities, and titers reported in this study may further support the applied goal of cost-effective, large -scale microbial lipid production for use as biofuels and biochemicals.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0492-3) contains supplementary material, which is available to authorized users.

Highlights

Microbial lipids are produced by many oleaginous organisms including the well-characterized yeast Yarrowia lipolytica, which can be engineered for increased lipid yield by up-regulation of the lipid biosynthetic pathway and down-regulation or deletion of competing pathways
Overexpression of endogenous DGA1 in Yarrowia lipolytica In an effort to develop a biocatalyst for industrial lipid production, we evaluated several Y. lipolytica wild-type strains for desirable biocatalyst qualities, such as minimal citric acid secretion, non-hyphal morphology, and ease of genetic manipulation
We confirmed that the overexpression of DGA1 increased lipid content in Y. lipolytica

Summary

Introduction

Microbial lipids are produced by many oleaginous organisms including the well-characterized yeast Yarrowia lipolytica, which can be engineered for increased lipid yield by up-regulation of the lipid biosynthetic pathway and down-regulation or deletion of competing pathways. Among natively oleaginous organisms, which are capable of producing lipids at a level greater than 20 % of their dry cell weight, the well-characterized yeast Yarrowia lipolytica [2] has received particular interest. Fatty acids are the dominant lipid compound, constituting 90–95 % of total lipid material [13, 14]. To avoid the potentially toxic and membrane-disturbing effects of free fatty acids, they are incorporated into nonpolar lipids such as triacylglycerols (TAG) and sterol esters. TAGs are stored in Friedlander et al Biotechnol Biofuels (2016) 9:77 subcellular compartments termed lipid droplets or lipid bodies [15,16,17,18]

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Conclusion