Affinity Purification of a Recombinant Protein Expressed as a Fusion with the Maltose-Binding Protein (MBP) Tag

Krisna C. Duong-Ly, Sandra B. Gabelli

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

Expression of fusion proteins such as MBP fusions can be used as a way to improve the solubility of the expressed protein in E. coli (Fox and Waugh, 2003; Nallamsetty et al., 2005; Nallamsetty and Waugh, 2006) and as a way to introduce an affinity purification tag. The protocol that follows was designed by the authors as a first step in the purification of a recombinant protein fused with MBP, using fast protein liquid chromatography (FPLC). Cells should have been thawed, resuspended in binding buffer, and lysed by sonication or microfluidization before mixing with the amylose resin or loading on the column. Slight modifications to this protocol may be made to accommodate both the protein of interest and the availability of equipment.

Original languageEnglish (US)
Title of host publicationMethods in Enzymology
PublisherAcademic Press Inc.
Pages17-26
Number of pages10
DOIs
StatePublished - Jun 20 2015

Publication series

NameMethods in Enzymology
Volume559
ISSN (Print)0076-6879
ISSN (Electronic)1557-7988

Keywords

  • Affinity chromatography
  • Affinity purification
  • Fast protein liquid chromatography (FPLC)
  • Fusion protein
  • MBP fusion
  • Maltose-binding protein (MBP)
  • Recombinant protein

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

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