Design and Engineering of Transmembrane Cytochromes

Abstract

Transmembrane electron transport (ET) is essential to the bioenergetic process that power life. The membrane-bound protein complexes that perform respiratory and photosynthetic ET have evolved over millions of years to be highly specialised and efficient but are difficult to engineer or use in synthetic biology. To overcome this complexity, we use computational protein design methods to build redox proteins from scratch. We use these proteins to build novel ET chains from the bottom up and to study the fundamental processes of ET.

I will first describe the design of our minimal artificial transmembrane cytochrome, CytbX. This is a simple membrane-bound four-helix bundle that binds two b-type heme molecules to facilitate transmembrane ET.  CytbX can be readily expressed in E. Coli and efficiently localises to the cell membrane where it binds heme in vivo. I will describe the redox properties of CytbX and how we have used it to study the phenomenon of redox cooperativity. We are now seeking to incorporate CytbX into larger functional ET complexes and develop the design principles to do so. I will share recent efforts to elongate CytbX to create extended heme wires, to design self-assembling CytbX oligomers, to incorporate CytbX into multi-protein ET complexes, and to equip CytbX with flavin cofactors for rudimentary light harvesting.