Split inteins are autoprocessing proteins composed of N- and C- terminal subunits, found in unicellular organisms across all domains of life. In these organisms, essential genes are split and under the regulation of two promoters. After translation the two ends come together and undergo protein splicing, removing the intein to form a functional protein. Inteins have been applied to advance drug development, gene therapy, structural determination, and regulation of function. However, for inteins to be used in these applications they must be accompanied by local, native amino acid sequences on either side of them to splice at optimal rates. The addition of a non-native sequence, at the splice site of a protein of interest, can have deleterious effects on protein structure and function. Thus, our goal was to determine the extein dependence of four fast-splicing, split inteins (gp41-1, gp41-8, IMPDH, and NrdJ-1) by using a kanamycin resistance assay. We constructed kanamycin resistance plasmids with varying extein scar lengths for each of our four split inteins. Moving forward, in addition to experimentally determining the extein dependence of selected split inteins, we hope to make inteins a more widely available tool for biochemical research by expanding our intein database that currently has over 2500 inteins logged.

Assembling split inteins for escape from kanamycin resistance protein

ACS Spring 2022

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.