Engineering the Stereoisomeric Structure of Plant Triacylglycerol to Mimic Human Milk Fat

Human milk fat substitute (HMFS) is a class of structured lipid that is widely used as an ingredient in infant formulas. Like human milk fat, HMFS is characterised by an enrichment of palmitoyl (C16:0) groups at the middle (sn-2 or β) position on the glycerol backbone, and there is evidence from multiple clinical trials that triacylglycerol (TAG) with this unusual stereoisomeric structure assists nutrient absorption in the neonatal gut. HMFS production currently relies on enzyme-based catalysis since there is no biological source of fat with the equivalent structure, other than humans. Around 1/2 a million tons of fat is used in infant formula production each year. The majority of this fat is obtained from plants, which specifically exclude C16:0 from the middle position. In this study we have engineered the metabolic pathway for TAG biosynthesis in the model oilseed Arabidopsis thaliana so that the percentage of C16:0 esterified to the middle (versus outer) position(s) is increased by more than 20-fold (i.e. from ~3% in wild type to >70% in our final iteration). We have also modified fatty acid biosynthesis so that total fatty acyl composition, as well as TAG structure, is more comparable to that of human milk fat. Finally, we have applied this technology to the oilseed crop Brassica napus and oleaginous yeast Yarrowia lipolytica. The work we will present therefore provides new biological sources of HMFS for infant formula.