Quantitative phosphoproteomic analysis of porcine muscle within 24 h postmortem

J Proteomics. 2014 Jun 25;106:125-39. doi: 10.1016/j.jprot.2014.04.020. Epub 2014 Apr 24.

Abstract

Protein phosphorylation can regulate most of the important processes in muscle, such as metabolism and contraction. The postmortem (PM) metabolism and rigor mortis have essential effects on meat quality. In order to identify and characterize the protein phosphorylation events involved in meat quality development, a quantitative mass spectrometry-based phosphoproteomic study was performed to analyze the porcine muscle within 24h PM using dimethyl labeling combined with the TiSH phosphopeptide enrichment strategy. In total 305 unique proteins were identified, including 160 phosphoproteins with 784 phosphorylation sites. Among these, 184 phosphorylation sites on 93 proteins had their phosphorylation levels significantly changed. The proteins involved in glucose metabolism and muscle contraction were the two largest clusters of phosphoproteins with significantly changed phosphorylation levels in muscle within 24 h PM. The high phosphorylation level of heat shock proteins (HSPs) in early PM may be an adaptive response to slaughter stress and protect muscle cell from apoptosis, as observed in the serine 84 of HSP27. This work indicated that PM muscle proteins underwent significant changes at the phosphorylation level but were relatively stable at the total protein level, suggesting that protein phosphorylation may have important roles in meat quality development through the regulation of proteins involved in glucose metabolism and muscle contraction, thereby affecting glycolysis and rigor mortis development in PM muscle.

Biological significance: The manuscript describes the characterization of postmortem (PM) porcine muscle within 24 h postmortem from the perspective of protein phosphorylation using advanced phosphoproteomic techniques. In the study, the authors employed the dimethyl labeling combined with the TiSH phosphopeptide enrichment and LC-MS/MS strategy. This was the first high-throughput quantitative phosphoproteomic study in PM muscle of farm animals. In the work, both the proteome and phosphoproteome were analyzed, and the large number of identified peptides, phosphopeptides and phosphorylation sites can greatly enrich the current farm animal protein database. The proteins involved in glycometabolism, muscle contraction and heat shock proteins (HSPs) showed significantly changed phosphorylation levels during PM meat development. This work indicated that PM muscle proteins underwent significant changes at phosphorylation level but were relatively stable at the total protein level, suggesting that protein phosphorylation may have important roles in meat development through the regulation of proteins involved in metabolism and muscle contraction, thereby affecting glycolysis and rigor mortis development in PM muscle. The work can promote the understanding of PM muscle metabolism and meat quality development, and be helpful for future meat quality control.

Keywords: Glucose metabolism; Meat quality development; Muscle contraction; Postmortem porcine muscle; Quantitative phosphoproteomics; Stress.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Motifs
  • Animals
  • Apoptosis
  • Chromatography, Liquid
  • Computational Biology
  • Databases, Protein
  • Glucose / metabolism
  • Glycolysis
  • HSP27 Heat-Shock Proteins / metabolism
  • Heat-Shock Proteins / metabolism
  • Hydrogen-Ion Concentration
  • Muscle Contraction
  • Muscles / metabolism*
  • Phosphoproteins / metabolism*
  • Phosphorylation
  • Protein Interaction Mapping
  • Proteomics / methods*
  • Rigor Mortis
  • Serine / metabolism
  • Swine
  • Tandem Mass Spectrometry
  • Trypsin / chemistry

Substances

  • HSP27 Heat-Shock Proteins
  • Heat-Shock Proteins
  • Phosphoproteins
  • Serine
  • Trypsin
  • Glucose