The Rainbow Graveyard

E. Coli is one of the gram-negative bacteria and most of their types are harmless. However some of E.Coli types such as enterohaemorrhagic E.Coli (EHEC) can cause serious diseases.

In our project we designed an innovative model to prevent gram-negative growth and infection. In our model, to prevent E.Coli growth, we modified non-infectious gram-positive bacteria B.Subtilis by transforming it with a construct, which was designed to produce limulus anti-LPS factor (LALF) together with a signal peptide. LALF is expected to bind and neutralize Lipopolysaccharide (LPS) found in gram-negative bacteria cell wall. On the other hand, to show E.Coli growth inhibition, we prepared another construct, which carries reflectin sequence with a signal peptide sequence and transferred into E.Coli. Reflectin protein produces color by reflecting light in different wavelenghts. This ability helps us to detect whether E.Coli is dead or alive...

We use two proteins which are anti-LPS (LALF) and Reflectin 1a. Anti-LPS protein can bind to LPS coat of gram negatives. With this function, the protein can be used as protection from septic shock. Also, anti LPS protein works as bacteriostatic. Reflectin 1a is a camouflage material and the protein is produced by many cephalopods (squid, cuttlefish, and octopus).
In this project, our first aim is to obtain an anti-gram negative bacterial surface with the help of anti-LPS protein. Our second aim is to sense the bacteriostatic effect with colorful reflectin protein. Also, we intend to apply our proteins on a surface as a coat.
We choose Bacillus Subtilis as the host of anti-LPS protein. Because, those bacteria are expected to produce biofilm containing the anti-LPS protein. This ability might help us to apply it on a surface.

To measure the inhibition effect properly, both of our bacteria are expected to synthesize reflectin. It is planned that anti-LPS exists in the biofilm will bind to LPS on the cell wall of Gram (-) bacteria, thus, Gram (-) bacteria will not grow and survive in our biofilm surface. Around the binding, reflectin thickness is going to decrease and change the color of the surface because of inhibition of growth of E.coli. So, if the color changes from red to blue, we can say there was a contact between E.coli and B.subtilis via anti-LPS factor. You can see for more information about LALF, Reflectin and Biofilm in the below links:

LALF (Anti-LPS)

Reflectin

Biofilm