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. 2014 Dec 19;3(12):969-71.
doi: 10.1021/sb500042m.

Engineered calcium-precipitable restriction enzyme

Josephina Hendrix  1 , Timothy Read, Jean-Francois Lalonde, Phillip K Jensen, William Heymann, Elijah Lovelace, Sarah A Zimmermann, Michael Brasino, Joseph Rokicki, Robin D Dowell
Affiliations

Engineered calcium-precipitable restriction enzyme

Josephina Hendrix et al. ACS Synth Biol. .

Abstract

We have developed a simple system for tagging and purifying proteins. Recent experiments have demonstrated that RTX (Repeat in Toxin) motifs from the adenylate cyclase toxin gene (CyaA) of B. pertussis undergo a conformational change upon binding calcium, resulting in precipitation of fused proteins and making this method a viable alternative for bioseparation. We have designed an iGEM Biobrick comprised of an RTX tag that can be easily fused to any protein of interest. In this paper, we detail the process of creating an RTX tagged version of the restriction enzyme EcoRI and describe a method for expression and purification of the functional enzyme.

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