Department of Biochemistry and Centre for Protein Science and Crystallography, The Chinese University of Hong Kong, Shatin, NT, Hong Kong, China.
Nucleic Acids Res. 2010 Oct;38(19):6803-12. doi: 10.1093/nar/gkq551. Epub 2010 Jun 17.
Maize ribosome-inactivating protein (RIP) is a plant toxin that inactivates eukaryotic ribosomes by depurinating a specific adenine residue at the α-sarcin/ricin loop of 28S rRNA. Maize RIP is first produced as a proenzyme with a 25-amino acid internal inactivation region on the protein surface. During germination, proteolytic removal of this internal inactivation region generates the active heterodimeric maize RIP with full N-glycosidase activity. This naturally occurring switch-on mechanism provides an opportunity for targeting the cytotoxin to pathogen-infected cells. Here, we report the addition of HIV-1 protease recognition sequences to the internal inactivation region and the activation of the maize RIP variants by HIV-1 protease in vitro and in HIV-infected cells. Among the variants generated, two were cleaved efficiently by HIV-1 protease. The HIV-1 protease-activated variants showed enhanced N-glycosidase activity in vivo as compared to their un-activated counterparts. They also possessed potent inhibitory effect on p24 antigen production in human T cells infected by two HIV-1 strains. This switch-on strategy for activating the enzymatic activity of maize RIP in target cells provides a platform for combating pathogens with a specific protease.
玉米核糖体失活蛋白（RIP）是一种植物毒素，通过脱嘌呤特定的腺嘌呤残基在 28S rRNA 的 α-亚精胺/蓖麻毒素环失活真核核糖体。玉米 RIP 首先作为一种前酶产生，在蛋白质表面有一个 25 个氨基酸的内部失活区。在萌发过程中，这种内部失活区的蛋白水解去除产生具有完整 N-糖苷酶活性的活性异源二聚体玉米 RIP。这种天然的开关机制为将细胞毒素靶向病原体感染的细胞提供了机会。在这里，我们报告了在内部失活区添加 HIV-1 蛋白酶识别序列，并在体外和感染 HIV 的细胞中通过 HIV-1 蛋白酶激活玉米 RIP 变体。在产生的变体中，有两种被 HIV-1 蛋白酶有效切割。与未激活的变体相比，HIV-1 蛋白酶激活的变体在体内显示出增强的 N-糖苷酶活性。它们还对两种 HIV-1 株感染的人 T 细胞中 p24 抗原的产生具有很强的抑制作用。这种在靶细胞中激活玉米 RIP 酶活性的开关策略为利用特定蛋白酶对抗病原体提供了一个平台。
