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Recent Advances in GFP Folding Reporter and Split-GFP Solubility Reporter Technologies. Application to Improving the Folding and Solubility of Recalcitrant Proteins from Mycobacterium tuberculosis

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Journal of Structural and Functional Genomics

Abstract

We have improved our green fluorescent protein (GFP) folding reporter technology [Waldo et al., (1999) Nat. Biotechnol. 17, 691–695] to evolve recalcitrant proteins from Mycobacterium tuberculosis. The target protein is inserted into the scaffolding of the GFP, eliminating false-positive artifacts caused by expression of truncated protein variants from internal cryptic ribosome binding sites in the target RNA. In parallel, we have developed a new quantitative fluorescent protein tagging and detection system based on micro-domains of GFP. This split-GFP system, which works both in vivo and in vitro, is amenable to high-throughput assays of protein expression and solubility [Cabantous et al., (2005) Nat. Biotechnol. 23, 102–107]. Together, the GFP folding reporter and split-GFP technologies offer a comprehensive system for manipulating and improving protein folding and solubility.

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Abbreviations

AnTET:

anhydrotetracycline

GFP:

green fluorescent protein

IPTG:

isopropyl thiogalactoside

MBP:

maltose-binding protein

NaCl:

sodium chloride

SUMO:

small ubiquitin-like modifiers

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Authors and Affiliations

  1. Bioscience Division, Los Alamos National Laboratory, MS-M888, Los Alamos, NM, 87545, USA

    Stéphanie Cabantous, Jean-Denis Pédelacq, Brian L. Mark, Cleo Naranjo, Thomas C. Terwilliger & Geoffrey S. Waldo

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  1. Stéphanie Cabantous
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  2. Jean-Denis Pédelacq
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  3. Brian L. Mark
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  4. Cleo Naranjo
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  5. Thomas C. Terwilliger
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  6. Geoffrey S. Waldo
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Correspondence to Thomas C. Terwilliger.

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Cabantous, S., Pédelacq, JD., Mark, B. et al. Recent Advances in GFP Folding Reporter and Split-GFP Solubility Reporter Technologies. Application to Improving the Folding and Solubility of Recalcitrant Proteins from Mycobacterium tuberculosis. J Struct Funct Genomics 6, 113–119 (2005). https://doi.org/10.1007/s10969-005-5247-5

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  • DOI: https://doi.org/10.1007/s10969-005-5247-5

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