The rational design and construction of a cuboidal iron–sulfur protein

Rational protein design is an emerging approach for testing general theories of protein chemistry through the creation of new structures and functions. Here we present the first successful introduction by rational design of a [Fe(4)S(4)] cuboidal cluster into the hydrophobic core of Escherichia coli thioredoxin, a protein normally devoid of metal centers. Cuboidal [Fe(4)S(4)] is one of the stable forms of self-assembled iron–sulfur clusters that are thought to represent some of the earliest evolved biological redox centers. [Fe(4)S(4)] clusters have been recruited for use in a variety of proteins whose functions are central to many of the major biochemical processes ranging from simple soluble electron-transfer agents, to membrane-bound components of electron-transfer chains, to electron reservoirs in complex metalloenzymes such as nitrogenase. By situating an [Fe(4)S(4)] cluster into a protein environment not previously adapted by evolution we can explore the factors by which their activity is modulated by the protein matrix.