Enhancing biofuel production through synthetic protein fusions

The development of lignocellulosic-based bioprocesses to make biofuels and other valuable chemical products is critically dependent on the efficiency with which bacteria can degrade, uptake and catabolise the sugars present in these feedstocks and then use the released energy and carbon skeleton to produce and export the desired products. While we know the transporters and enzymes used by bacteria to access some of the components of lignocellulose, such as the sugar xylose, no attempt has been made to engineer these existing pathways to enhance the rate at which catabolism occurs. We aim to achieve this by using a novel and exciting method that this based on the concept of spatial coupling of related reactions in the cell. This is to physically scaffold the proteins involved in a particular catabolic pathway into an assembly, reducing the time for diffusion of products from one reaction to be substrate for the next. This concept has recently been used successfully to increase the flux through the threonine biosynthetic pathway in E. coli by fusing the enzymes to zinc-finger proteins that then bind to a DNA scaffold (Lee et al ., 2012 Applied Environmental Microbiol 79:774-782). However, in this synthetic biology project we will primarily investigate a novel route to spatial coupling using genetically encoded rod-like protein domains, recently discovered in York in the lab of Prof. Jennifer Potts, to bring together sets of functionally related proteins. These rod-like domains can be made to a variety of different lengths to control the distance between interacting proteins. Uniquely we will also attempt to incorporate membrane transporters into the complex to more closely couple uptake to catabolism. This timely project also has the potential to apply the same approach to other pathways relevant for biofuel and chemical production.