Part:BBa_K5035004:Design

A0A419DFF0 is a corrinoid iron-sulfur-containing reductive dehalogenase produced by the Dehalococcoidia bacterium. A0A419DFF0 is structurally similar to enzyme A6RdhA, a corrinoid iron-sulfur-containing reductive dehalogenase capable of degrading PFOA. Because of A0A419DFF0's structural and familial similarity to A6RdhA, it is predicted to have similar PFOA degrading abilities.

A0A419DFF0 requires sulfur, iron, and norpseudo vitamin B12 to fold properly. The 2024 IEA iGEM team successfully expressed A0A419DFF0 within MyTXTL and incorporated the necessary cofactors within its structure using the cofactor reconstitution protocol detailed by Nakamura et al. The Results of the team's expression of A0A419DFF0 are shown in the gel below.

Protein gel electrophoresis of purified A0A419DFF0 solution shows a band at the expected size of around 46 kDa.

Design Notes

A hexahistidine tag for nickel column purification was added to A0A419DFF0.

References

https://www.uniprot.org/uniprotkb/A0A419DFF0/entry

Nakamura, R., Obata, T., Nojima, R., Hashimoto, Y., Noguchi, K., Ogawa, T., & Yohda, M. (2018). Functional Expression and Characterization of Tetrachloroethene Dehalogenase From Geobacter sp. Frontiers in microbiology, 9, 1774. https://doi.org/10.3389/fmicb.2018.01774