Exploring different strategies to purify Vibrio cholerae ParE2

Scriptie/Masterproef: Master's Thesis

Samenvatting

In both archaea and bacteria, the small operons of toxin-antitoxin (TA) systems are omnipresent. They have been suggested to contribute to the stabilization of mobile genetic elements, bacteriophage protection, stress response and more controversial, persister formation. One of such TA systems is parDE2 from Vibrio cholerae. The toxin, ParE2, targets DNA Gyrase, a class II topoisomerase which translates itself as a mechanism of interest in novel antibiotics development. A major bottleneck opposing the in vitro study of the toxin, is its inherent cytotoxicity, making it troublesome to purify at high quantities. This study therefore investigates four different production strategies to ameliorate the production numbers, without interfering with the activity of the toxin. Of the four strategies, two provide the most promising results, rooted in two different organisms, namely bacterial and insect cells. This article demonstrates that recombinantly expressed VcParE2 variants are correctly folded and retain their functionality. Furthermore, the described insect cell protocol offers a five-fold higher yield via an intronic expression system, compared to a bacterial unfolding-refolding protocol.
Datum prijs12 sep 2022
Originele taalEnglish
Prijsuitreikende instantie
  • Biologie
  • Vrije Universiteit Brussel
BegeleiderRemy Loris (Promotor) & Yana Girardin (Advisor)

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