College of Life Sciences, State Key Laboratory of Crop Genetics and Germplasm Enhancement, Nanjing Agricultural University, Nanjing, 210095, PR China.
Jilin Provincial Key Laboratory of Agricultural Biotechnology, Jilin Academy of Agricultural Sciences, Changchun, 130033, PR China.
Biochem Biophys Res Commun. 2021 Sep 3;568:143-150. doi: 10.1016/j.bbrc.2021.06.087. Epub 2021 Jun 30.
Triacylglycerol (TAG), a main component of oil, is mainly biosynthesized by diacylglycerol acyltransferase (DGAT), which is critical for oil accumulation in plants. Intensive focus has been on DGAT2 functioning in unsaturated fatty acids biosynthesis. In this study, we analyzed the coding sequence (CDS) and amino acid sequence of GmDGAT2A and determined its key active sites through site-directed mutagenesis. As a consequence, H132, G201, and P152-X-I154-K155 were found to be essential active sites for GmDGAT2A. The spatial structure of the protein may bring the three active sites into close proximity, constituting an active domain. Additionally, N-terminus of GmDGAT2A was found to be an important regulator for the activity. Further, in vitro activity results uncovered GmDGAT2A was prone to utilize C18:2-CoA as the substrate. Consequently, overexpression of GmDGAT2A driven by a seed-specific promoter of Gmole1 in soybean significantly increased linoleic acid content specifically and total oil content, concomitant with accelerated elongation.
甘油三酯（TAG）是油的主要成分，主要由二酰基甘油酰基转移酶（DGAT）合成，这对于植物中的油脂积累至关重要。人们一直高度关注 DGAT2 在不饱和脂肪酸生物合成中的作用。在这项研究中，我们分析了 GmDGAT2A 的编码序列（CDS）和氨基酸序列，并通过定点突变确定了其关键活性位点。结果发现，H132、G201 和 P152-X-I154-K155 是 GmDGAT2A 的必需活性位点。该蛋白质的空间结构可能使三个活性位点接近，构成一个活性域。此外，还发现 GmDGAT2A 的 N 端是其活性的重要调节剂。此外，体外活性结果表明，GmDGAT2A 更倾向于利用 C18:2-CoA 作为底物。因此，在大豆中，由 Gmole1 种子特异性启动子驱动的 GmDGAT2A 的过表达特异性地显著增加了亚油酸含量和总油含量，同时加速了伸长。
