Preparation of recombinant peptides with site- and degree-specific lysine (13)C-methylation

Biochemistry. 2009 May 12;48(18):3798-800. doi: 10.1021/bi900348z.

Abstract

Lysine methylation is an important post-translational modification that affects protein function; for example, the transcriptional activity of the p53 tumor suppressor protein. To facilitate structural characterization of complexes involving proteins and methylated targets by nuclear magnetic resonance spectroscopy, we devised a simple method for preparing recombinant (15)N/(13)C-enriched peptides with a (13)C-methyl-labeled methylated lysine analogue. The method, which relies on the synthesis of (13)C-enriched alkylating agents, was applied to the production of 15-residue p53 peptides variously methylated at lysine analogue 370. The peptides were used to probe the methylation state-dependent interactions of mono, di, and trimethylated p53 with three different proteins.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Carbon Isotopes / chemistry*
  • Chromatography, Liquid
  • Lysine / metabolism*
  • Methylation
  • Nuclear Magnetic Resonance, Biomolecular
  • Peptides / isolation & purification
  • Peptides / metabolism*
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism

Substances

  • Carbon Isotopes
  • Peptides
  • Recombinant Proteins
  • Lysine