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Methods Mol Biol. 2016;1436:109-18. doi: 10.1007/978-1-4939-3667-0_8.

Large-Scale Overproduction and Purification of Recombinant Histone Deacetylase 8 (HDAC8) from the Human-Pathogenic Flatworm Schistosoma mansoni.

Author information

1
Département de Biologie Structurale Intégrative, Institut de Génétique et Biologie Moléculaire et Cellulaire (IGBMC), Université de Strasbourg (UDS), CNRS, INSERM, 1 rue Laurent Fries, B.P. 10142, 67404, Illkirch Cedex, France.
2
Center for Infection and Immunity of Lille (CIIL), INSERM U1019 - CNRS UMR 8204, Université de Lille, Institut Pasteur de Lille, 1 rue Professeur Calmette, 59019, Lille Cedex, France.
3
Département de Biologie Structurale Intégrative, Institut de Génétique et Biologie Moléculaire et Cellulaire (IGBMC), Université de Strasbourg (UDS), CNRS, INSERM, 1 rue Laurent Fries, B.P. 10142, 67404, Illkirch Cedex, France. romier@igbmc.fr.

Abstract

Epigenetic mechanisms underlie the morphological transformations and shifts in virulence of eukaryotic pathogens. The targeting of epigenetics-driven cellular programs thus represents an Achilles' heel of human parasites. Today, zinc-dependent histone deacetylases (HDACs) belong to the most explored epigenetic drug targets in eukaryotic parasites. Here, we describe an optimized protocol for the large-scale overproduction and purification of recombinant smHDAC8, an emerging epigenetic drug target in the multicellular human-pathogenic flatworm Schistosoma mansoni. The strategy employs the robustness of recombinant expression in Escherichia coli together with initial purification through a poly-histidine affinity tag that can be removed by the thrombin protease. This protocol is divided into two steps: (1) large-scale production of smHDAC8 in E. coli, and (2) purification of the target smHDAC8 protein through multiple purification steps.

KEYWORDS:

Enzyme; Histone deacetylase; Purification; Recombinant expression; Schistosoma

PMID:
27246211
DOI:
10.1007/978-1-4939-3667-0_8
[Indexed for MEDLINE]

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