Affinity purification of gst fusion proteins for immunohistochemical studies of gene expression

Melania Mercado-Pimentel, Nicole C. Jordan, Gabriel O. Aisemberg

Research output: Contribution to journalArticle

18 Scopus citations

Abstract

Fusion proteins expressed in bacteria are often insoluble or inefficiently purified by standard procedures previously reported to work well for the non-fused proteins. We report here a simple but general procedure that can be used to quickly customize and optimize the purification of milligram quantities of most GST fusion proteins. For each new protein, this procedure determines the optimal conditions for solubilization with detergents in a bacterial lysate, binding to glutathione-agarose beads, and elution with different buffers. This approach was applied to three GST fusion proteins containing large fragments of the Hox transcription factors Lox2, Lox4, and Lox6 that had low solubility and poor elution when purified following published procedures. After optimization, purified proteins were obtained at high yield and successfully used to raise and purify antibodies for the study of the expression patterns of these genes in embryonic tissues.

Original languageEnglish (US)
Pages (from-to)260-265
Number of pages6
JournalProtein Expression and Purification
Volume26
Issue number2
DOIs
Publication statusPublished - 2002
Externally publishedYes

    Fingerprint

ASJC Scopus subject areas

  • Biochemistry

Cite this