Background

The voltage output part of our project aims to mimic the signal transduction that occurs at a neural synapse. We are engineering E.coli to create a voltage output on detection of glutamate. This imitates the creation of a postsynaptic potential in a dendrite when a neurotransmitter (such as glutamate) is present at the synapse. The mechanism we have designed is similar to that used in the brain – relying on ion movement across the membrane, and gated ion channels. To simplify the concept, we are only regulating and measuring the flux of potassium (K+) ions, and we are using a directly glutamate-gated K+ ion channel. This means that on the binding of glutamate, the channels will open, allowing a K+ flux, which will change the voltage of the medium enough to be detected with a very sensitive electrode. In order to set up a large enough K+ concentration gradient across the membrane for ions to flow down when the channels open, an ion pump is necessary. E.coli has a transmembrane P-type ATPase called Kdp, which pumps K+ into the cell. We have isolated this gene to overexpress it, therefore causing the cells to pump in a large number of K+ ions. However, E.coli also has a number of osmoregulatory systems which use relative K+ ion concentrations to control turgor. There are K+ leak channels (Kch and Kef) in the membrane, so we have ordered E.coli strains with mutations in these genes to allow K+ to remain sequestered inside the cells until the glutamate-gated channels open.

Presentation