PDF(Pubmed)
Abstract：
Plant legumains are crucial for processing seed storage proteins and are critical regulators of plant programmed cell death. Although research on legumains boosted recently, little is known about their activity regulation. In our study, we used pull-down experiments to identify AtCYT6 as a natural inhibitor of legumain isoform β (AtLEGβ) in Arabidopsis thaliana. Biochemical analysis revealed that AtCYT6 inhibits both AtLEGβ and papain-like cysteine proteases through two separate cystatin domains. The N-terminal domain inhibits papain-like proteases, while the C-terminal domain inhibits AtLEGβ. Furthermore, we showed that AtCYT6 interacts with legumain in a substrate-like manner, facilitated by a conserved asparagine residue in its reactive center loop. Complex formation was additionally stabilized by charged exosite interactions, contributing to pH-dependent inhibition. Processing of AtCYT6 by AtLEGβ suggests a context-specific regulatory mechanism with implications for plant physiology, development, and programmed cell death. These findings enhance our understanding of AtLEGβ regulation and its broader physiological significance.
摘要：
植物豆科植物对于处理种子储存蛋白至关重要，并且是植物程序性细胞死亡的关键调节剂。尽管最近对豆类的研究有所增加，对他们的活动调节知之甚少。在我们的研究中,我们使用下拉实验来鉴定AtCYT6是拟南芥中豆科蛋白酶同工型β（AtLEGβ）的天然抑制剂。生化分析表明，AtCYT6通过两个独立的胱抑素结构域抑制AtLEGβ和木瓜蛋白酶样半胱氨酸蛋白酶。N端结构域抑制木瓜蛋白酶样蛋白酶，而C端结构域抑制AtLEGβ。此外，我们发现AtCYT6与legumain以类似底物的方式相互作用，由其反应中心环中保守的天冬酰胺残基促进。通过带电的外部位相互作用额外稳定了复合物的形成，有助于pH依赖性抑制。AtLEGβ对AtCYT6的加工提示了对植物生理学有影响的上下文特异性调节机制。发展,和程序性细胞死亡。这些发现增强了我们对AtLEGβ调节及其更广泛的生理意义的理解。
