Figure 5.
The muscle mitochondrial acetylproteome is largely unaffected by NR treatment
(A) Proteomics workflow. Quadriceps muscle tissue was obtained from HF- and HF + NR-fed FC mice. Proteins were extracted, enzymatically digested, labeled with unique tandem mass tag (TMT) 10plex reagents, and pooled. After retaining a small portion of the input fraction, acetyl-peptides were enriched via immunoprecipitation, and the flow-through was subsequently used for phosphoenrichment via IMAC. After analyzing all fractions by nanoflow liquid chromatography-tandem mass spectrometry (nLC-MS/MS), the quantitative data was analyzed using Proteome Discoverer 2.4 (PD 2.4) and in-house Python code.
(B) Volcano plot of acetyl-peptide relative occupancy (protein-normalized acetylation) versus −log (p value), comparing FC mice with and without NR supplementation. Open and closed dots refer to non-mitochondrial and mitochondrial peptides, respectively, with size inversely correlated with quantitative FDR. N = 5 per group. N represents biological replicates.