Abstract：
Selenocysteine (Sec) is the 21st proteogenic amino acid and it is now widely accepted that Sec is involved in redox biochemistry when incorporated in proteins. However, many of the chemical mechanisms for Sec bioactivity remain unknown. Herein, we describe a derivative of Sec, alpha-methyl Sec ((αMe)Sec), that is a useful chemical tool to study selenoenzyme mechanisms. (αMe)Sec is identical to Sec except the Cα-H is replaced with a Cα-methyl group, which prevents this derivative from undergoing oxygen-mediated β-syn elimination to dehydroalanine, which is a common problem with Sec-containing peptides and proteins. Thus, since (αMe)Sec-containing peptides and proteins cannot lose the side-chain selenium atom when oxidized, mechanistic studies can be performed that are not always possible with Sec. In this chapter, we provide detailed methods for the incorporation of (αMe)Sec into peptides using solid phase peptide synthesis and subsequent incorporation into mammalian thioredoxin reductase using protein semisynthesis. We then provide two examples of how (αMe)Sec has been used as a chemical tool to study selenoenzyme mechanism. Finally, we discuss future applications where we envision (αMe)Sec will be useful.
摘要：
硒代半胱氨酸（Sec）是第21种蛋白质氨基酸，现在被广泛接受的是，当将Sec掺入蛋白质中时，它参与氧化还原生物化学。然而,Sec生物活性的许多化学机制仍然未知。在这里，我们描述了Sec的导数，α-甲基Sec((αMe)Sec),这是研究硒酶机制的有用化学工具。（αMe）Sec与Sec相同，除了Cα-H被Cα-甲基取代，防止该衍生物经历氧介导的β-syn消除脱氢丙氨酸，这是含有Sec的肽和蛋白质的常见问题。因此，因为含（αMe）Sec的肽和蛋白质在氧化时不能失去侧链硒原子，可以进行机械研究，这在SEC中并不总是可行的。在这一章中,我们提供了使用固相肽合成将（αMe）Sec掺入肽中并随后使用蛋白质半合成掺入哺乳动物硫氧还蛋白还原酶中的详细方法。然后，我们提供了两个示例，说明如何将（αMe）Sec用作研究硒酶机制的化学工具。最后,我们讨论了未来的应用，我们设想(αMe)Sec将是有用的。
