NMR-based screening of proteins containing 13C-labeled methyl groups

Philip J. Hajduk, David J. Augeri, Jamey Mack, Renaldo Mendoza, Jianguo Yang, Stephen F. Betz, Stephen W. Fesik

Research output: Contribution to journalArticlepeer-review

188 Scopus citations

Abstract

A method is described for NMR-based screening that involves monitoring the 13C/1H chemical shift changes of a protein selectively labeled with 13C at the methyl groups of valine, leucine, and isoleucine (δ1 only). Using this approach, the sensitivity is increased by nearly 3-fold compared with that of NMR-based screening using 1H/15N chemical shifts. A synthetic route is described for the inexpensive production of the labeled amino acid precursors [3,3'-13C]-α-ketoisovalerate and [3-13C]-α-ketobutyrate, making the cost of protein preparation comparable to that of uniform 15N labeling. In addition to enhancing the NMR-based screening efforts directed against low molecular weight proteins (MW ≤ 30 kDa), the use of the selective methyl labels in combination with deuterium labeling is advantageous for screening high molecular weight protein targets (MW ≥ 100 kDa).

Original languageEnglish (US)
Pages (from-to)7898-7904
Number of pages7
JournalJournal of the American Chemical Society
Volume122
Issue number33
DOIs
StatePublished - Aug 23 2000
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry

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