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 disease.

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 gram negatives’ LPS coat and with this function the protein can use protecting from septic shock. Also anti LPS protein works bacteriostatic. And 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 obtain an anti-gram negative bacterial surface with the help of anti-LPS protein. Our second aim is sensing the bacteriostatic effect with colorful reflectin protein. Also we intend to apply our proteins on a surface as a coat.
We use Bacillus Subtilis as a host because those bacteria are expected to produce biofilm containing the anti-LPS protein that can be applied on a surface and this might be the best coating material for our project. Also, by killing spores of B.subtilis on the biofilm surface with aqueous dissolved oxygen, ascorbic acid, and copper ions we can protect from some patogene effects of B.subtilis. and we use e. coli as gram negative bacteria.
Both of our bacteria are expected to synthesize reflectin. When the biofilm containing anti-LPS factor (LALF) from B.subtilis, it will bind to LPS on Gram (-) bacteria’s cell wall and Gram (-) bacteria will not grow and survive in our biofilm surface. Around the binding, reflectin thickness is going to decrease and change the surface colours because of E.coli’s death. So if the colour 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

LALF (Anti-LPS)

Reflectin

Biofilm