Sayanova Olga, Haslam Richard, Qi Baoxiu, Lazarus Colin M, Napier Johnathan A
Rothamsted Research, Harpenden, Herts AL5 2JQ, UK.
FEBS Lett. 2006 Apr 3;580(8):1946-52. doi: 10.1016/j.febslet.2006.02.062. Epub 2006 Mar 3.
A cDNA encoding a C20 Delta8-desaturase was isolated from the free-living soil amoeba, Acanthamoeba castellanii and functionally characterised by heterologous expression. The open reading frame of the A. castellanii C20 Delta8-desaturase showed similarity to other microsomal front-end desaturases, but the N-terminal domain contained a variant form of the conserved heme-binding motif in which H-P-G-G is replaced by H-P-A-G. Co-expression of the A. castellani Delta8-desaturase with the Isochrysis galbana Delta9-elongase in transgenic Arabidopsis plants confirmed the activity observed in yeast and its role in the alternative pathway for C20 polyunsaturated fatty acid synthesis. Acyl-CoA profiles of these transgenic plants revealed an unexpected accumulation of C20 fatty acids in the acyl-CoA pool. This is the first report of an alternative pathway C20 Delta8-desaturase from a non-photosynthetic organism, and also the first report of a front-end desaturase lacking the canonical cytochrome b5 domain.
从自由生活的土壤变形虫——卡氏棘阿米巴中分离出一个编码C20 Δ8-去饱和酶的cDNA，并通过异源表达对其进行了功能表征。卡氏棘阿米巴C20 Δ8-去饱和酶的开放阅读框与其他微粒体前端去饱和酶具有相似性，但其N端结构域包含保守的血红素结合基序的变体形式，其中H-P-G-G被H-P-A-G取代。在转基因拟南芥植物中将卡氏棘阿米巴Δ8-去饱和酶与等鞭金藻Δ9-延长酶共表达，证实了在酵母中观察到的活性及其在C20多不饱和脂肪酸合成的替代途径中的作用。这些转基因植物的酰基辅酶A谱显示，酰基辅酶A池中C20脂肪酸意外积累。这是关于来自非光合生物的替代途径C20 Δ8-去饱和酶的首次报道，也是关于缺乏典型细胞色素b5结构域的前端去饱和酶的首次报道。
