Department of Horticultural Sciences, University of Florida, Gainesville, Florida, 32611, USA.
Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Ann Arbor, Michigan, 48109, USA.
Plant J. 2018 Jul;95(2):358-370. doi: 10.1111/tpj.13955. Epub 2018 Jun 12.
The proteinogenic branched-chain amino acids (BCAAs) leucine, isoleucine and valine are essential nutrients for mammals. In plants, BCAAs double as alternative energy sources when carbohydrates become limiting, the catabolism of BCAAs providing electrons to the respiratory chain and intermediates to the tricarboxylic acid cycle. Yet, the actual architecture of the degradation pathways of BCAAs is not well understood. In this study, gene network modeling in Arabidopsis and rice, and plant-prokaryote comparative genomics detected candidates for 3-methylglutaconyl-CoA hydratase (4.2.1.18), one of the missing plant enzymes of leucine catabolism. Alignments of these protein candidates sampled from various spermatophytes revealed non-homologous N-terminal extensions that are lacking in their bacterial counterparts, and green fluorescent protein-fusion experiments demonstrated that the Arabidopsis protein, product of gene At4g16800, is targeted to mitochondria. Recombinant At4g16800 catalyzed the dehydration of 3-hydroxymethylglutaryl-CoA into 3-methylglutaconyl-CoA, and displayed kinetic features similar to those of its prokaryotic homolog. When at4g16800 knockout plants were subjected to dark-induced carbon starvation, their rosette leaves displayed accelerated senescence as compared with control plants, and this phenotype was paralleled by a marked increase in the accumulation of free and total leucine, isoleucine and valine. The seeds of the at4g16800 mutant showed a similar accumulation of free BCAAs. These data suggest that 3-methylglutaconyl-CoA hydratase is not solely involved in the degradation of leucine, but is also a significant contributor to that of isoleucine and valine. Furthermore, evidence is shown that unlike the situation observed in Trypanosomatidae, leucine catabolism does not contribute to the formation of the terpenoid precursor mevalonate.
蛋白质支链氨基酸（BCAAs）亮氨酸、异亮氨酸和缬氨酸是哺乳动物的必需营养物质。在植物中，当碳水化合物受到限制时，BCAAs 可作为替代能源，BCAA 的分解代谢为呼吸链提供电子和三羧酸循环的中间产物。然而，BCAA 降解途径的实际结构尚未得到很好的理解。在这项研究中，通过拟南芥和水稻的基因网络建模以及植物-原核生物比较基因组学，检测到 3-甲基戊二酰辅酶 A 水合酶（4.2.1.18）的候选基因，该酶是亮氨酸分解代谢中缺失的植物酶之一。从各种有胚植物中取样的这些蛋白质候选物的比对揭示了缺乏其细菌对应物的非同源性 N 端延伸，绿色荧光蛋白融合实验表明，拟南芥蛋白，At4g16800 的产物，靶向线粒体。重组 At4g16800 催化 3-羟甲基戊二酰辅酶 A 脱水生成 3-甲基戊二酰辅酶 A，并显示出与其原核同源物相似的动力学特征。当 at4g16800 敲除植物受到黑暗诱导的碳饥饿时，与对照植物相比，它们的莲座叶显示出加速衰老的现象，并且这种表型与游离和总亮氨酸、异亮氨酸和缬氨酸的显著积累相平行。突变体 at4g16800 的种子也表现出类似的游离 BCAAs 积累。这些数据表明，3-甲基戊二酰辅酶 A 水合酶不仅参与亮氨酸的降解，而且也是异亮氨酸和缬氨酸降解的重要贡献者。此外，有证据表明，与锥虫科观察到的情况不同，亮氨酸分解代谢不会导致萜类前体甲羟戊酸的形成。
