Abstract
Thraustochytrids possess docosahexaenoic acid (DHA, 22:6n-3) as acyl chain(s) of triacylglycerol (TG) and phosphatidylcholine (PC), some of which contain multiple DHAs. However, little is known about how these DHA-rich glycerolipids are produced in thraustochytrids. In this study, we identified PLAT2 in Aurantiochytrium limacinum F26-b as a glycerol-3-phosphate (G3P) acyltransferase (GPAT) by heterologous expression of the gene in budding yeast. Subsequently, we found that GPAT activity was reduced by disruption of the PLAT2 gene in A. limacinum, resulting in a decrease in DHA-containing lysophosphatidic acid (LPA 22:6). Conversely, overexpression of PLAT2 increased both GPAT activity and LPA 22:6. These results indicate that PLAT2 is a GPAT that transfers DHA to G3P in vivo as well as in vitro. Overexpression of the PLAT2 gene increased the production of a two DHA-containing diacylglycerol (DG 44:12), followed by an increase in the three DHA-containing TG (TG 66:18), two-DHA-containing TG (TG 60:12), and two DHA-containing PC (PC 44:12). However, overexpression of PLAT2 did not increase DHA-free DG (DG32:0), which was preferentially converted to three 16:0-containing TG (TG 48:0) but not two 16:0-containing PC (PC 32:0). Collectively, we revealed that DHA-rich glycerolipids are produced from a precursor, LPA 22:6, which is generated by incorporating DHA to G3P by PLAT2 in the A. limacinum.
Highlights
N-3 polyunsaturated fatty acids (n-3 PUFAs), such as docosahexaenoic acid (DHA, 22:6n-3), decrease the level of blood neutral fat and function in preventing arteriosclerosis and thrombus formation, thereby reducing the risk of cardiovascular diseases [1, 2]
We found several putative gene products as lysophospholipid acyltransferase (LPLAT) homologues in A. limacinum, which we tentatively designated as PLAT1~7 in this study
Based on the phylogenetic tree, all PLATs found in thraustochytrids were in the AGPAT family, and PhosphoLipid AcylTransferase 2 (PLAT2) was selected as a candidate for glycerol-3-phosphate acyltransferase (GPAT) because it was evolutionary close to mammalian and Drosophila GPATs
Summary
N-3 polyunsaturated fatty acids (n-3 PUFAs), such as docosahexaenoic acid (DHA, 22:6n-3), decrease the level of blood neutral fat and function in preventing arteriosclerosis and thrombus formation, thereby reducing the risk of cardiovascular diseases [1, 2]. An enzyme for producing DHA-rich glycerolipids in thraustochytrids
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