Journal of Biomolecular NMR

, Volume 11, Issue 1, pp 97–102

An efficient and cost-effective isotope labeling protocol for proteins expressed in shape Escherichia coli

Authors

  • Mengli Cai
    • Laboratory of Chemical Physics
  • Ying Huang
    • Laboratory of Molecular BiologyNational Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health
  • Kazuyasu Sakaguchi
    • Laboratory of Cell Biology, Building 37National Cancer Institute, National Institutes of Health
  • G. Marius Clore
    • Laboratory of Chemical Physics
  • Angela M. Gronenborn
    • Laboratory of Chemical Physics
  • Robert Craigie
    • Laboratory of Chemical Physics
Article

DOI: 10.1023/A:1008222131470

Cite this article as:
Cai, M., Huang, Y., Sakaguchi, K. et al. J Biomol NMR (1998) 11: 97. doi:10.1023/A:1008222131470

Abstract

A cost-effective protocol for uniform 15N and/or13 C isotope labeling of bacterially expressed proteins is presented. Unlike most standard protocols, cells are initially grown in a medium containing nutrients at natural abundance and isotopically labeled nutrients are only supplied at the later stages of growth and during protein expression. This permits the accumulation of a large cell mass without the need to employ expensive isotopically labeled nutrients. The abrupt decrease in oxygen consumption that occurs upon complete exhaustion of essential nutrients is used to precisely time the switch between unlabeled and labeled nutrients. Application of the protocol is demonstrated for wild-type and a mutant of the N-terminal zinc-binding domain of HIV-1 integrase.

heteronuclear NMRisotope labelingprotein expression
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Copyright information

© Kluwer Academic Publishers 1998