title: Metabolic engineering Corynebacterium glutamicum to produce triacylglycerols.
abstract: In this study, we metabolically engineered Corynebacterium glutamicum to produce triacylglycerols (TAGs) by completing and constraining a de novo TAG biosynthesis pathway. First, the plasmid pZ8_TAG4 was constructed which allows the heterologous expression of four genes: three (atf1 and atf2, encoding the diacylglycerol acyltransferase; pgpB, encoding the phosphatidic acid phosphatase) to complete the TAG biosynthesis pathway, and one gene (tadA) for lipid body assembly. Second, we applied four metabolic strategies to increase TAGs accumulation: (i) boosting precursor supply by heterologous expression of tesA (encoding thioesterase to form free fatty acid to reduce the feedback inhibition by acyl-ACP) and fadD (encoding acyl-CoA synthetase to enhance acyl-CoA supply), (ii) reduction of TAG degradation and precursor consumption by deleting four cellular lipases (cg0109, cg0110, cg1676 and cg1320) and the diacylglycerol kinase (cg2849), (iii) enhancement of fatty acid biosynthesis by deletion of fasR (cg2737, TetR-type transcriptional regulator of genes for the fatty acid biosynthesis), and (iv) elimination of the observed by-product formation of organic acids by blocking the acetic acid (pqo) and lactic acid production (ldh) pathways. The final strain (CgTesRtcEfasEbp/pZ8_TAG4) achieved a 7.5% yield of total fatty acids (2.38 ± 0.05 g/L intracellular fatty acids and 0.64 ± 0.09 g/L extracellular fatty acids) from 4% glucose in shake flasks after process optimization. This corresponds to maximum intracellular fatty acids content of 17.8 ± 0.5% of the dry cell.
authors.affiliation: Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Jens
authors.initials: J
authors.lastname: Plassmeier
authors.affiliation: Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, PR China.
authors.forename: Youyuan
authors.initials: Y
authors.lastname: Li
authors.affiliation: Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Christian
authors.initials: C
authors.lastname: Rueckert
authors.affiliation: Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Engineering Systems Division, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Anthony J
authors.initials: AJ
authors.lastname: Sinskey
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D048230:Corynebacterium glutamicum / Q000235:genetics* / Q000378:metabolism*; D024861:Genetic Enhancement / Q000379:methods; D060847:Metabolic Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D014280:Triglycerides / Q000096:biosynthesis* / Q000302:isolation & purification
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; D014280:Triglycerides
keywords: Corynebacterium glutamicum; Metabolic engineering; Microbial lipids; TAG biosynthesis; Triacylglycerol