Karchi H, Shaul O, Galili G
Department of Plant Genetics, Weizmann Institute of Science, Rehovot, Israel.
Proc Natl Acad Sci U S A. 1994 Mar 29;91(7):2577-81. doi: 10.1073/pnas.91.7.2577.
The regulation of synthesis and accumulation of the essential amino acid lysine was studied in seeds of transgenic tobacco plants expressing, in a seed-specific manner, two feedback-insensitive bacterial enzymes: dihydrodipicolinate synthase (EC 4.2.1.52) and aspartate kinase (EC 2.7.2.4). High-level expression of the two bacterial enzymes resulted in only a slight increase in free lysine accumulation at intermediate stages of seed development, while free lysine declined to the low level of control plants toward maturity. To test whether enhanced catabolism may have contributed to the failure of free lysine to accumulate in seeds of transgenic plants, we analyzed the activity of lysine-ketoglutarate reductase (EC 1.5.1.7), an enzyme that catabolizes lysine into saccharopine. In both the control and the transgenic plants, the timing of appearance of lysine-ketoglutarate reductase activity correlated very closely with that of dihydrodipicolinate synthase activity, suggesting that lysine synthesis and catabolism were coordinately regulated during seed development. Notably, the activity of lysine-ketoglutarate reductase was significantly higher in seeds of the transgenic plants than in the controls. Coexpression of both bacterial enzymes in the same plant resulted in a significant increase in the proportions of lysine and threonine in seed albumins. Apparently, the normal low steady-state levels of free lysine and threonine in tobacco seeds may be rate limiting for the synthesis of seed proteins, which are relatively rich in these amino acids.
在以种子特异性方式表达两种反馈不敏感细菌酶（二氢二吡啶二羧酸合酶（EC 4.2.1.52）和天冬氨酸激酶（EC 2.7.2.4））的转基因烟草植物种子中，研究了必需氨基酸赖氨酸的合成和积累调控。这两种细菌酶的高水平表达在种子发育的中间阶段仅导致游离赖氨酸积累略有增加，而游离赖氨酸在接近成熟时降至对照植物的低水平。为了测试增强的分解代谢是否可能导致转基因植物种子中游离赖氨酸积累失败，我们分析了赖氨酸-酮戊二酸还原酶（EC 1.5.1.7）的活性，该酶将赖氨酸分解代谢为saccharopine。在对照植物和转基因植物中，赖氨酸-酮戊二酸还原酶活性出现的时间与二氢二吡啶二羧酸合酶活性的时间非常密切相关，这表明种子发育过程中赖氨酸的合成和分解代谢受到协调调控。值得注意的是，转基因植物种子中赖氨酸-酮戊二酸还原酶的活性显著高于对照。在同一植物中共表达这两种细菌酶导致种子白蛋白中赖氨酸和苏氨酸的比例显著增加。显然，烟草种子中游离赖氨酸和苏氨酸正常的低稳态水平可能是种子蛋白质合成的限速因素，种子蛋白质相对富含这些氨基酸。
