Team:Kent
Welcome to our iGem page
Our project aims to provide an alternative solution to produce nano wire. Our system takes advantage of the endogenous curli system of E.coli to produce functional extracellular amyloid composed of sup35. The curli system uses the Sec transport pathway to translocate sup35 through the inner membrane where it is transported from the periplasm through a specific pore to the outer membrane where it assembles into repeating units. We intend to engineer the sup35 to have a bound cytochrome b562 that will transport electrons along the amyloid fibrils. Due to export relying on the Sec pathway cytochrome b562 cannot properly fold because it requires the cofactor haem, which we are able to add to the growth medium to facilitate folding.
The wider aim of the project is to funnel electrons from the electron transport chain into the sup35/cytochrome b562 nanowire which would provide a completely renewable source of energy that could be used to power small consumer products.
How we’re going to do it
We are going to utilise the endogenous curli system of E.coli VS45 strain to export a protein that will form amyloid fibres. The protein will be expressed from a plasmid that we are going to engineer with the sup35 and cytochrome b562 gene inserted into the expression site.
The curli system consists of csgG export system which recognises the csgA N-terminal signal sequence that is attached to the sup35 protein. This will allow the sup35/cytochrome b562 complex to be translocated to the outer membrane to form an amyloid fibre.
To reach the curli-specific pore on the outer membrane the sup35/cytochrome b562 complex must pass the inner membrane to the periplasm via the Sec translocation pathway. Cytochrome b562 requires the cofactor heam to allow correct folding. Cofactors cannot bind before translocation via the Sec pathway, however cytochrome b562 can bind to haem that is exogenously added to the solution after export, thus allowing the cytochrome to fold into its active conformation. The cytochrome on the amyloid fibres will then transport electrons down the transport chain, acting as a nanowire.
We would create the nanowire by forming a biofilm over a surface that we would like electricity to flow across. This could be used as a replacement for conventional nanowire allowing the downscaling of consumer products. The ultimate goal of the project is to funnel electrons from the electron transport chain of the bacterial respiratory chain into the amyloid chain. This would allow the production of a self-powering unit that aggregates its own nanowire.