Fig. 3: Liver-specific nascent proteome identification and analysis in living mice.

a Nascent protein incorporated by AlkK containing an alkyne handle could be labeled by azide-PEG4-Biotin, and then be enriched by streptavidin. The reaction conditions, including extraction method, concentration of Cu²⁺ and ligand, and reaction time, are optimized to improve the labeling efficiency in the liver extracts. b Labeling and enrichment of the nascent proteins of the whole proteome by western blot. SA-Blot, immunoblotting with streptavidin-HRP. Experiment was independently repeated three times. c Overlap of proteins identified between the control group and SORT-AC group. The proteins that were detected only in the SORT-AC groups were highlighted as SORT-AC Unique. d Volcano plot of the proteins identified in both the control group and SORT-AC group. The dashed lines were under the significance curve parameter of S₀ = 0.1 and a false discovery rate <0.05. The proteins significantly enriched were colored in blue and highlighted as SORT-AC Enrich. Statistical significance was assessed using a two-sided, empirical Bayes moderated t-test. e The strategy of identifying SORT-AC Print proteins. f Tissue specific analysis with previously reported dataset among total expressed proteome, total identified proteins, SORT-AC Print, respectively. g Enrichment analysis of SORT-AC Print proteins that major participate pathways with a one-sided Fisher’s exact test. The blue indicated the pathways highly related to liver functions. The resulting p-values were adjusted for multiple hypothesis testing using the Benjamini–Hochberg procedure. h The interaction network analysis of SORT-AC Print proteins. The membrane proteins were colored in green. i The ratio of membrane and transmembrane proteins in SORT-AC Print (top) and the annotation of the subcellular localization of SORT-AC Print membrane proteins (bottom). j Overlap of membrane proteins in SORT-AC Print in mitochondrion, endoplasmic reticulum, and Golgi apparatus. k Overlap of transmembrane proteins in SORT-AC Print in mitochondrion, endoplasmic reticulum, and Golgi apparatus. l Annotation of transmembrane proteins involved in lipid metabolism in SORT-AC Print, which were located in mitochondrion and endoplasmic reticulum. Source data are provided as a Source Data file.