Proteomic analysis of organ-specific post-translational lysine-acetylation and -methylation in mice by use of anti-acetyllysine and -methyllysine mouse monoclonal antibodies

Proteomics. 2005 Dec;5(18):4653-64. doi: 10.1002/pmic.200500042.

Abstract

Post-translational lysine-acetylation and -methylation are two major PTMs of lysine residues in proteins. Recently, we established pan-reactive anti-acetyllysine mouse mAbs, which can bind to Nepsilon-acetylated lysine residues in various contexts of amino acid sequences. In the present study, we established pan-reactive anti-methyllysine mouse mAbs comparable to the anti-acetyllysine ones. By using these anti-acetyllysine and -methyllysine antibodies, we found that the pattern of lysine-acetylated and -methylated proteins in mouse organs showed extreme variation from organ to organ. We selected brain and skeletal muscle as model cases to be further analyzed by 2-DE followed by Western blotting. In brain, alpha-tubulin at its basal level was found to be extremely acetylated; and alpha-enolase was shown to be a newly recognized possibly acetylated protein. NF-L protein, Hsc70, alpha-tubulin fragments, beta-actin, and brain-type creatine kinase were identified as putative lysine-methylated proteins in mouse brain. In skeletal muscle, lysine-methylation of alpha-actin and both lysine-acetylation and -methylation of muscle-type creatine kinase were found as novel putative lysine-modified proteins. The approach presented here might be useful to find novel disease markers and/or drug target molecules that would not be noticed by use of the traditional proteomic approach only.

MeSH terms

  • Acetylation
  • Actins / metabolism
  • Animals
  • Antibodies, Monoclonal / biosynthesis
  • Brain Chemistry
  • Creatine Kinase, MM Form / analysis
  • Cross Reactions
  • Female
  • Histones / metabolism
  • Horseradish Peroxidase / immunology
  • Hybridomas / immunology
  • Lysine / analogs & derivatives*
  • Lysine / immunology
  • Lysine / metabolism*
  • Methylation
  • Mice
  • Muscle Proteins / chemistry
  • Muscle, Skeletal / chemistry
  • Nerve Tissue Proteins / chemistry
  • Organ Specificity
  • Protein Processing, Post-Translational / genetics*
  • Proteomics*
  • Tubulin / metabolism

Substances

  • Actins
  • Antibodies, Monoclonal
  • Histones
  • Muscle Proteins
  • Nerve Tissue Proteins
  • Tubulin
  • epsilon-N-methyllysine
  • N(alpha)-acetyllysine
  • 2-methyllysine
  • N-epsilon-acetyllysine
  • Horseradish Peroxidase
  • Creatine Kinase, MM Form
  • Lysine