Marsh E N G
University of Michigan, Ann Arbor, MI, United States.
Methods Enzymol. 2016;580:251-78. doi: 10.1016/bs.mie.2016.05.006. Epub 2016 Jun 24.
As methods to incorporate noncanonical amino acid residues into proteins have become more powerful, interest in their use to modify the physical and biological properties of proteins and enzymes has increased. This chapter discusses the use of highly fluorinated analogs of hydrophobic amino acids, for example, hexafluoroleucine, in protein design. In particular, fluorinated residues have proven to be generally effective in increasing the thermodynamic stability of proteins. The chapter provides an overview of the different fluorinated amino acids that have been used in protein design and the various methods available for producing fluorinated proteins. It discusses model proteins systems into which highly fluorinated amino acids have been introduced and the reasons why fluorinated residues are generally stabilizing, with particular reference to thermodynamic and structural studies from our laboratory. Lastly, details of the methodology we have developed to measure the thermodynamic stability of oligomeric fluorinated proteins are presented, as this may be generally applicable to many proteins.
随着将非天然氨基酸残基掺入蛋白质的方法变得更加强大，人们对利用它们来改变蛋白质和酶的物理及生物学特性的兴趣与日俱增。本章讨论了在蛋白质设计中使用疏水性氨基酸的高度氟化类似物，例如六氟亮氨酸。特别是，氟化残基已被证明在提高蛋白质的热力学稳定性方面通常是有效的。本章概述了已用于蛋白质设计的不同氟化氨基酸以及用于生产氟化蛋白质的各种可用方法。它讨论了已引入高度氟化氨基酸的模型蛋白质系统以及氟化残基通常具有稳定作用的原因，特别参考了我们实验室的热力学和结构研究。最后，介绍了我们开发的用于测量寡聚氟化蛋白质热力学稳定性的方法细节，因为这可能普遍适用于许多蛋白质。
