Falco S C, Guida T, Locke M, Mauvais J, Sanders C, Ward R T, Webber P
E. I. DuPont de Nemours & Co., Wilmington, DE 19880-0402, USA.
Biotechnology (N Y). 1995 Jun;13(6):577-82. doi: 10.1038/nbt0695-577.
We have increased the lysine content in the seeds of canola and soybean plants by circumventing the normal feedback regulation of two enzymes of the biosynthetic pathway, aspartokinase (AK) and dihydrodipicolinic acid synthase (DHDPS). Lysine-feedback-insensitive bacterial DHDPS and AK enzymes encoded by the Corynebacterium dapA gene and a mutant E. coli lysC gene, respectively, were linked to a chloroplast transit peptide and expressed from a seed-specific promoter in transgenic canola and soybean seeds. Expression of Corynebacterium DHDPS resulted in more than a 100-fold increase in the accumulation of free lysine in the seeds of canola; total seed lysine content approximately doubled. Expression of Corynebacterium DHDPS plus lysine-insensitive E. coli AK in soybean transformants similarly caused several hundred-fold increases in free lysine and increased total sed lysine content by as much as 5-fold. Accumulation of alpha-amino adipic acid (AA) in canola and saccharopine in soybean, which are intermediates in lysine catabolism, was also observed.
我们通过规避生物合成途径中两种酶——天冬氨酸激酶（AK）和二氢吡啶二羧酸合酶（DHDPS）的正常反馈调节，提高了油菜籽和大豆种子中的赖氨酸含量。分别由棒状杆菌dapA基因和突变型大肠杆菌lysC基因编码的对赖氨酸反馈不敏感的细菌DHDPS和AK酶，与叶绿体转运肽相连，并在转基因油菜籽和大豆种子中由种子特异性启动子表达。棒状杆菌DHDPS的表达导致油菜籽种子中游离赖氨酸的积累增加了100多倍；种子总赖氨酸含量大约翻了一番。在大豆转化体中表达棒状杆菌DHDPS加对赖氨酸不敏感的大肠杆菌AK同样导致游离赖氨酸增加数百倍，种子总赖氨酸含量增加多达5倍。还观察到油菜籽中α-氨基己二酸（AA）和大豆中saccharopine的积累，它们是赖氨酸分解代谢的中间产物。
