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Rapid analysis of protein expression and solubility with the SpyTag-SpyCatcher system

Dovala, Dustin and Sawyer, William and Rath, Christopher and Metzger Iv, Louis (2016) Rapid analysis of protein expression and solubility with the SpyTag-SpyCatcher system. Protein Expr Purif, 117. pp. 44-51. ISSN 1096-0279 (Electronic); 1046-5928 (Linking)

Abstract

Successful isolation of well-folded and active protein often first requires the creation of many constructs. These are needed to assess the effects of truncations, insertions, mutations, and the presence and position of different affinity tags. Determining which constructs yield the highest expression and solubility requires the investigator to express and partially purify each construct, and, in the case of low-expressing proteins, to follow the protein using time-consuming Western blots. Even then, many proteins form soluble aggregates, which may only be apparent after more extensive purification via size exclusion chromatography. In this work, we have utilized a covalent bond-forming tag/domain pair, known as SpyTag/SpyCatcher, to rapidly and specifically attach a fluorescent label to proteins of interest in cellular lysates. Once labeled, tagged proteins can easily be followed via SDS-PAGE and fluorescence size exclusion chromatography (F-SEC) to assess expression levels, solubility, and monodispersity without the need for purification. These techniques enable rapid and facile analysis of proteins, which may greatly facilitate optimization of protein expression constructs.

Item Type: Article
Additional Information: Unmapped bibliographic data: LA - eng [Field not mapped to EPrints] ET - 2015/09/26 [Field not mapped to EPrints] ST - Rapid analysis of protein expression and solubility with the SpyTag-SpyCatcher system [Field not mapped to EPrints] AU - Dovala, D. [Field not mapped to EPrints] AU - Sawyer, W. S. [Field not mapped to EPrints] AU - Rath, C. M. [Field not mapped to EPrints] AU - Metzger, L. E. th [Field not mapped to EPrints] AD - Novartis Institutes for BioMedical Research, Emeryville, CA, United States. [Field not mapped to EPrints] DP - NLM [Field not mapped to EPrints] AN - 26405011 [Field not mapped to EPrints]
Keywords: Dispersity
Date Deposited: 14 Dec 2016 00:45
Last Modified: 14 Dec 2016 00:45
URI: https://oak.novartis.com/id/eprint/26851

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