Difference between revisions of "Part:BBa K4586013"

Revision as of 17:43, 3 October 2023

CX43

Part Description

This part is a transmembrane protein called connexin (CX43) that oligomerizes to form hexameric channels, called hemichannels, that can control the interaction and transfer of information between exosomes and cells of interest, which is called intercellular communication.

Usage

This part is implicated in our design to increase the amount of our therapeutic agent within the exosomes, as the CX43 transmembrane portion facilitates the loading of our cargo into the exosomes.

Literature Characterization

The study made western blot analyses of extracts from various cell lines to find out the effect of connexin 43 in different cells.

By using Connexin 43 (E7N2R) XP Rabbit mAb in the upper and GAPDH (D16H11) XP Rabbit mAb in the lower. Connexin 43 protein is expressed at low levels in BeWo cells, which is compatible with the expected expression pattern.

In lane 1, we have Western blot analysis of extracts from control 293T cells; in lane 2, we have Western blot analysis of connexin 43 knockout 293T cells, and that was done using Connexin 43 XP Rabbit mAb in the upper and GAPDH XP Rabbit mAb in the lower. Connexin 43 antibody specificity is demonstrated by the absence of a signal in connexin 43 knockout 293T cells.

Characterization By Mutational Landscape

In order to optimize the function of our parts, we've used the concept of Directed Evolution through applying different mutations and measuring the effects of these mutations on their evolutionary epistatic fitness. As displayed in the chart below, the mutation (V236I) shows the highest epistatic fitness, while the lowest score was associated with the mutation (K109Q,S244R).

Figure . An illustration of the effects of different mutations on the Epistatic Fitness of CX43.

References

Giepmans, B.N. et al. (2001) J Biol Chem 276, 8544-9.

Sequence and Features