Team:Queens Canada/Lab Overview

Overview

Here we designed a fluorescent anti-THC antibody, optimized for E. coli expression. Recombinant antibody expression in E. coli is notoriously challenging, as typical IgG proteins require post-translational modifications. However, Recombinant expression of antibodies can be made possible by truncating antibody fragment (Fig. 1). The protein sequence for the anti-THC fragment (ScFv) had been previously characterized and is optimized for E. coli expression systems (1). Hence, this part is codon optimized for E. coli. The protein sequence also contains a periplasmic localization signal (PLS), encoded by the OmpA sequence. Periplasmic localization provides an oxidizing environment for disulfide bonds in the antibody to form. to further improve disulfide bond formation, the protein may be expressed in an engineered cell line with an oxidizing cytoplasmic environment (Ex: Rosetta gami2).

Figure 1. The modification of an IgG protein to a Fab and ScFv, which can be expressed in E. coli. In the case of the anti-THC antibody, the protein was conjugated to a fluorescent protein at the C-terminal.

References

1. Brennan, J. (2005) The production of recombinant single chain antibody fragments for the detection of illicit drug residues. doctoral thesis, Dublin City University, [online] http://doras.dcu.ie/17319/ (Accessed March 12, 2019)