Abstract
BackgroundMonounsaturated fatty acids (MUFAs) are the best components for biodiesel when considering the low temperature fluidity and oxidative stability. However, biodiesel derived from vegetable oils or microbial lipids always consists of significant amounts of polyunsaturated and saturated fatty acids (SFAs) alkyl esters, which hampers its practical applications. Therefore, the fatty acid composition should be modified to increase MUFA contents as well as enhancing oil and lipid production.ResultsThe model microorganism Escherichia coli was engineered to produce free MUFAs. The fatty acyl-ACP thioesterase (AtFatA) and fatty acid desaturase (SSI2) from Arabidopsis thaliana were heterologously expressed in E. coli BL21 star(DE3) to specifically release free unsaturated fatty acids (UFAs) and convert SFAs to UFAs. In addition, the endogenous fadD gene (encoding acyl-CoA synthetase) was disrupted to block fatty acid catabolism while the native acetyl-CoA carboxylase (ACCase) was overexpressed to increase the malonyl coenzyme A (malonyl-CoA) pool and boost fatty acid biosynthesis. The finally engineered strain BL21ΔfadD/pE-AtFatAssi2&pA-acc produced 82.6 mg/L free fatty acids (FFAs) under shake-flask conditions and FFAs yield on glucose reached about 3.3% of the theoretical yield. Two types of MUFAs, palmitoleate (16:1Δ9) and cis-vaccenate (18:1Δ11) made up more than 75% of the FFA profiles. Fed-batch fermentation of this strain further enhanced FFAs production to a titer of 1.27 g/L without affecting fatty acid compositions.ConclusionsThis study demonstrated the possibility to regulate fatty acid composition by using metabolic engineering approaches. FFAs produced by the recombinant E. coli strain consisted of high-level MUFAs and biodiesel manufactured from these fatty acids would be more suitable for current diesel engines.
Highlights
Monounsaturated fatty acids (MUFAs) are the best components for biodiesel when considering the low temperature fluidity and oxidative stability
Production of free unsaturated fatty acids by a specific thioesterase As a model organism to study fatty-acid biosynthesis, E. coli has a type II fatty acid synthase (FAS) (FAS II) system, which is found in most bacteria and plants [27]
These two enzymes can cleave the bonds of fatty acyl-acyl carrier protein (ACP) thioesters, the catalytic efficiency is much lower than acyl-coenzyme A (CoA) esters of the same length [30]
Summary
Monounsaturated fatty acids (MUFAs) are the best components for biodiesel when considering the low temperature fluidity and oxidative stability. Escherichia coli possesses many advantages over other microorganisms, for example, a clear genetic background, the convenience of its ability to be genetically modified, and good growth properties with low nutrient requirements [12,13] It has been widely studied for production of fatty acids [14,15], or directly biodiesel [16,17], and shows a promising prospect for industrial application. The irreversible synthesis of malonyl-CoA catalyzed by acetyl-CoA carboxylase is another key regulatory point for fatty acid production Overexpression of this enzyme could increase the final titer of FFAs in the engineered strain [20]. Modifying the fatty acid compositions of E. coli is critical to promote its practical application in the biodiesel industry
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