Rational design of nascent metalloenzymes

Understanding the early genesis of new enzymatic functions is one of the ch allenges in protein design, mechanistic enzymology, and molecular evolution . We have experimentally mimicked starting points in this process by introd ucing primitive iron and oxygen binding sites at various locations in thior edoxin, a small protein lacking metal centers, by using computational desig n. These rudimentary active sites show emerging enzymatic activities that s elect to varying degrees between different oxygen chemistries. Even within these nascent enzymes, mechanisms by which different reactions are controll ed can be discerned. These involve both stabilizing and destabilizing inter actions imposed on the metal center by the surrounding protein matrix.