Rathore Ray Singh, Mishra Manjari, Pareek Ashwani, Singla-Pareek Sneh Lata
Plant Stress Biology, International Centre for Genetic Engineering and Biotechnology, New Delhi, 110067, India.
Stress Physiology and Molecular Biology Laboratory, School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India.
J Exp Bot. 2025 Mar 13;76(5):1408-1426. doi: 10.1093/jxb/erae414.
Amino acids are a major source of nourishment for people living in regions where rice is a staple food. However, rice grain is deficient in essential amino acids including lysine. The activity of the enzyme dihydrodipicolinate synthase (DHDPS) is crucial for lysine production in higher plants, but it is tightly regulated through feedback inhibition by its end product, lysine, leading to limited activity in the grain and resulting in low lysine accumulation. We identified lysine binding sites in the DHDPS enzyme and introduced key mutations to make DHDPS lysine feedback insensitive. Using in vivo analysis and functional complementation assays, we confirmed that protein engineering of the DHDPS renders it insensitive to lysine. Expression of mutated DHDPS resulted in 29% higher lysine and 15% higher protein accumulation in rice grains than in the wild type. Importantly, the lysine content in transgenic grains was maintained in cooked rice. The transgenic plants also exhibited enhanced stress tolerance along with higher antioxidant levels, improved photosynthesis, and higher grain yield compared to wild-type plants. We have shown that protein engineering of DHDPS in rice can lead to accumulation of lysine in grains and impart abiotic stress tolerance. This approach could improve health in regions with nutrient deficiencies and environmental stressors that challenge food production and human health.
氨基酸是生活在以大米为主食地区的人们的主要营养来源。然而，稻谷缺乏包括赖氨酸在内的必需氨基酸。二氢吡啶二羧酸合酶（DHDPS）的活性对高等植物中赖氨酸的产生至关重要，但它受到其终产物赖氨酸的反馈抑制的严格调控，导致在谷粒中的活性有限，从而导致赖氨酸积累量低。我们在DHDPS酶中鉴定出赖氨酸结合位点，并引入关键突变以使DHDPS对赖氨酸反馈不敏感。通过体内分析和功能互补试验，我们证实了DHDPS的蛋白质工程使其对赖氨酸不敏感。与野生型相比，突变型DHDPS的表达使水稻谷粒中的赖氨酸含量提高了29%，蛋白质积累量提高了15%。重要的是，转基因谷粒中的赖氨酸含量在煮熟的米饭中得以保持。与野生型植物相比，转基因植物还表现出更强的胁迫耐受性、更高的抗氧化水平、改善的光合作用和更高的谷物产量。我们已经表明，水稻中DHDPS的蛋白质工程可以导致谷粒中赖氨酸的积累，并赋予非生物胁迫耐受性。这种方法可以改善营养缺乏和环境压力挑战粮食生产和人类健康的地区的健康状况。
