Zhang Y, Yüksel K U, Gracy R W
Department of Biochemistry and Molecular Biology, University of North Texas Health Science Center, Fort Worth 76107.
Arch Biochem Biophys. 1995 Feb 20;317(1):112-20. doi: 10.1006/abbi.1995.1142.
Triosephosphate isomerase (TPI) provides an excellent model for terminal marking and protein degradation. Mammalian TPI is terminally modified by deamidation at Asn71-Gly, resulting in unfolding, dissociation, and proteolysis. In contrast, chicken TPI, which contains a lysine at position 71, is terminally modified by the oxidation of Cys126. Thus, both deamidation and oxidation initiate degradation of TPI from different species. To explore the terminal marking in other avians, we have purified the turkey TPI to homogeneity and determined its characteristics. Although the molecular properties of the turkey and chicken TPI were very similar, their tolerances to temperature, oxidants, and alkaline pH were very different. For example, chicken TPI was inactivated 80% in either 10 mM oxidized glutathione or H2O2, whereas 120 mM GSSG had no effect on turkey TPI, and > 120 mM H2O2 was needed for 80% inactivation. Under alkaline conditions that cause rapid deamidation of the mammalian TPI, neither avian enzyme deamidated. Chicken TPI, however, aggregated. Aggregation was reversed by 2-mercaptoethanol. Under prolonged exposure to milder conditions the turkey enzyme was completely inactivated and deamidated at Asn15-Gly. Thus, there are marked differences in the susceptibility of these two avian enzymes to oxidation and deamidation, and their terminal marking mechanisms appear to be different.
磷酸丙糖异构酶（TPI）为末端标记和蛋白质降解提供了一个绝佳的模型。哺乳动物的TPI在Asn71 - Gly处通过脱酰胺作用进行末端修饰，导致其解折叠、解离并被蛋白酶水解。相比之下，鸡的TPI在第71位含有赖氨酸，其通过Cys126的氧化进行末端修饰。因此，脱酰胺作用和氧化作用均引发了不同物种TPI的降解。为了探究其他鸟类中的末端标记情况，我们已将火鸡TPI纯化至同质，并确定了其特性。尽管火鸡和鸡的TPI分子特性非常相似，但它们对温度、氧化剂和碱性pH的耐受性却大不相同。例如，鸡的TPI在10 mM氧化型谷胱甘肽或H2O2中会有80%失活，而120 mM的谷胱甘肽二硫化物对火鸡TPI没有影响，且需要> 120 mM的H2O2才能使火鸡TPI有80%失活。在导致哺乳动物TPI快速脱酰胺的碱性条件下，两种鸟类的酶均不会脱酰胺。然而，鸡的TPI会聚集。2 - 巯基乙醇可使聚集逆转。在长时间暴露于较温和的条件下，火鸡的酶会完全失活，并在Asn15 - Gly处脱酰胺。因此，这两种鸟类的酶对氧化和脱酰胺的敏感性存在显著差异，并且它们的末端标记机制似乎也不同。
