IBBMC, CNRS UMR 8619, Univ Paris-Sud, Orsay, France.
Proteomics. 2010 Jul;10(13):2418-28. doi: 10.1002/pmic.200900835.
The importance of redox-regulation in Arabidopsis thaliana roots has been investigated through the identification of the proteins interacting with thioredoxin (TRX), an ubiquitous thiol-disulfide reductase. We have applied a proteomic approach based on affinity chromatography on a monocysteinic mutant of plastidial y-type TRX used as a bait to trap putative partners in a crude extract of root proteins. Seventy-two proteins have been identified, functioning mainly in metabolism, detoxification and response to stress, protein processing and signal transduction. This study allowed us to isolate 24 putative new targets and to propose the mevalonic acid-dependent biosynthesis of isoprenoids as a new redox-mediated process. The redox-regulation of phenylpropanoid biosynthesis is also suggested, three enzymes of this pathway being retained on the column. We also provided experimental evidence that phenylammonia-lyase was enzymatically more active when reduced by TRXy in root crude extract. Among the high number of partners involved in defense against stress we isolated from the column, we focused on plastidial monodehydroascorbate reductase and showed that its activity was dramatically increased in vitro in the presence of DTT-reduced TRXy1 in root crude extracts. Our data strongly suggest that TRXy1 could be the physiological regulator of monodehydroascorbate reductase in root plastids.
已通过鉴定与硫氧还蛋白（TRX）相互作用的蛋白质来研究拟南芥根中的氧化还原调节的重要性，TRX 是一种普遍存在的硫醇-二硫键还原酶。我们应用了一种基于质体 y 型 TRX 单半胱氨酸突变体的亲和层析的蛋白质组学方法，将其作为诱饵来捕获根蛋白粗提物中的假定伙伴。已经鉴定出 72 种蛋白质，主要功能是代谢、解毒和应激反应、蛋白质加工和信号转导。这项研究使我们能够分离出 24 种假定的新靶标，并提出依赖于甲羟戊酸的类异戊二烯生物合成是一种新的氧化还原介导的过程。苯丙烷生物合成的氧化还原调节也被提出，该途径的三种酶保留在柱子上。我们还提供了实验证据，表明在根粗提物中，TRXy 通过还原苯氨甲酰裂解酶使其酶活性增加。我们从柱子上分离出大量参与应激防御的伙伴，其中我们重点研究了质体单脱氢抗坏血酸还原酶，并表明在根粗提物中存在 DTT 还原的 TRXy1 时，其体外活性显著增加。我们的数据强烈表明，TRXy1 可能是根质体中单脱氢抗坏血酸还原酶的生理调节剂。
