Over-expression and purification strategies for recombinant multi-protein oligomers: A case study of Mycobacterium tuberculosis sigma/anti-sigma factor protein complexes

Thakur, Krishan Gopal and Jaiswal, Ravi Kumar and Shukla, Jinal K and Praveena, T and Gopal, B (2010) Over-expression and purification strategies for recombinant multi-protein oligomers: A case study of Mycobacterium tuberculosis sigma/anti-sigma factor protein complexes. In: Protein Expression and Purification, 74 (2). pp. 223-230.

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Abstract

The function of a protein in a cell often involves coordinated interactions with one or several regulatory partners. It is thus imperative to characterize a protein both in isolation as well as in the context of its complex with an interacting partner. High resolution structural information determined by X-ray crystallography and Nuclear Magnetic Resonance offer the best route to characterize protein complexes. These techniques, however, require highly purified and homogenous protein samples at high concentration. This requirement often presents a major hurdle for structural studies. Here we present a strategy based on co-expression and co-purification to obtain recombinant multi-protein complexes in the quantity and concentration range that can enable hitherto intractable structural projects. The feasibility of this strategy was examined using the sigma factor/anti-sigma factor protein complexes from Mycobacterium tuberculosis. The approach was successful across a wide range of sigma factors and their cognate interacting partners. It thus appears likely that the analysis of these complexes based on variations in expression constructs and procedures for the purification and characterization of these recombinant protein samples would be widely applicable for other multi-protein systems. (C) 2010 Elsevier Inc. All rights reserved.

Item Type:	Journal Article
Publication:	Protein Expression and Purification
Publisher:	Elsevier Science
Additional Information:	Copyright of this article belongs to Elsevier Science.
Keywords:	Sigma factor; Anti-sigma factor; Co-expression; Mycobacterium tuberculosis; Recombinant; Protein complex
Department/Centre:	Division of Biological Sciences > Molecular Biophysics Unit
Date Deposited:	13 Nov 2010 10:23
Last Modified:	13 Nov 2010 10:23
URI:	http://eprints.iisc.ac.in/id/eprint/33643

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