Expression and purification of Src-family kinases for solution NMR studies

Andrea Piserchio, David Cowburn, Ranajeet Ghose

Research output: Chapter in Book/Report/Conference proceedingChapter

3 Citations (Scopus)

Abstract

NMR analyses of the structure, dynamics, and interactions of the Src family kinases (SFKs) have been hindered by the limited ability to obtain sufficient amounts of properly folded, soluble protein from bacterial expression systems, to allow these studies to be performed in an economically viable manner. In this chapter, we detail our attempts to overcome these difficulties using the catalytic domain (SrcCD) of c-Src, the prototypical SFK, as an illustrative example. We describe in detail two general methods to express and purify SrcCD from Escherichia coli expression systems in both fully active wild-type and kinase-deficient mutant forms, allowing the efficient and cost-effective labeling by NMR-active isotopes for solution NMR studies.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
Pages111-131
Number of pages21
Volume831
DOIs
StatePublished - 2012

Publication series

NameMethods in Molecular Biology
Volume831
ISSN (Print)10643745

Fingerprint

src-Family Kinases
Bacterial Proteins
Isotopes
Catalytic Domain
Phosphotransferases
Escherichia coli
Costs and Cost Analysis

Keywords

  • Escherichia coli expression systems
  • Protein tyrosine kinases
  • Src-family kinases

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

Cite this

Piserchio, A., Cowburn, D., & Ghose, R. (2012). Expression and purification of Src-family kinases for solution NMR studies. In Methods in Molecular Biology (Vol. 831, pp. 111-131). (Methods in Molecular Biology; Vol. 831). https://doi.org/10.1007/978-1-61779-480-3_7

Expression and purification of Src-family kinases for solution NMR studies. / Piserchio, Andrea; Cowburn, David; Ghose, Ranajeet.

Methods in Molecular Biology. Vol. 831 2012. p. 111-131 (Methods in Molecular Biology; Vol. 831).

Research output: Chapter in Book/Report/Conference proceedingChapter

Piserchio, A, Cowburn, D & Ghose, R 2012, Expression and purification of Src-family kinases for solution NMR studies. in Methods in Molecular Biology. vol. 831, Methods in Molecular Biology, vol. 831, pp. 111-131. https://doi.org/10.1007/978-1-61779-480-3_7
Piserchio A, Cowburn D, Ghose R. Expression and purification of Src-family kinases for solution NMR studies. In Methods in Molecular Biology. Vol. 831. 2012. p. 111-131. (Methods in Molecular Biology). https://doi.org/10.1007/978-1-61779-480-3_7
Piserchio, Andrea ; Cowburn, David ; Ghose, Ranajeet. / Expression and purification of Src-family kinases for solution NMR studies. Methods in Molecular Biology. Vol. 831 2012. pp. 111-131 (Methods in Molecular Biology).
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