Team:US AFRL CarrollHS/Improve


BBa_K1321340, a double cellulose binding domain with an N-terminal linker, from Plate 5, Well 18A of the 2017 iGEM Distribution Kit was used by our team to develop a new and beneficial part. The part links the csgA gene from curli fiber DNA to the double cellulose binding domain (dCBD) gene. This new part has been submitted and labelled as BBa_K2522000.

Curli fibers are strands of proteins produced by certain strains of E. coli . The fibers are constructed on the outer surface of the cell membrane and encase the bacteria in an extracellular matrix. The genes that make up the DNA of curli fibers are referred to as csg (curli specific gene), and then a letter of the alphabet. Specifically, csgA functions as the building blocks of the hair-like portion of the fibers. The double cellulose binding domain, as the name suggests, attaches to cellulose when cellulose is present.

To construct the part, the csgA gene was extracted from the genomic DNA of E. coli Nissle and the dCBD was extracted from the iGEM Distribution Kit. The stop codon of the csgA was removed and the two genes were ligated together. In this manner, every time the csgA gene is expressed, a double cellulose binding domain would also be expressed so that the two parts function as a single unit.

Once a strain of curli-producing E. coli accepts a plasmid containing the csgA-dCBD part, the bacteria can encapsulate itself in cellulose. Such a feature can prove beneficial in various situations. In the context of our project, the cellulose would preserve the engineered microbe from being broken down too early when ingested, or protect the microbe if placed into drinking water. The part could be useful in similar applications where bacteria requires a protective feature. Consequently, developing a plasmid containing csgA-dCBD has improved upon the original function of the double cellulose binding domain part from the Distribution Kit.