Figure 4.

Limited mixing and exchange of mt-LLPs. (A) Schematic illustrating DSSO cross-linking of mitochondria isolated from ¹⁵N-pulse–labeled heart tissue extracts, sample processing, and MS1/MS2/MS3 analysis. (B) Silver-stained gel of control and cross-linked heart mitochondrial isolations. DSSO was added at a concentration of 1 mM; control was treated with DMSO only. Overview of cross-links between different mitochondrial proteins identified in the heart. (C) Number of cross-links identified in heart mitochondria according to the cross-link type. (D) Number of cross-links identified in heart mitochondria according to the peptide isotope build. (E) Map of the mitochondrial protein cross-links. ¹⁵N-¹⁵N (new-new) peptide cross-links are shown in blue; ¹⁴N-¹⁴N (old-old) peptide cross-links are shown in yellow; mixed ¹⁵N-¹⁴N (new-old) peptide cross-links are shown in green. (F) Summary of identified cross-linked peptides broken down by cross-link type and nitrogen isotype. Maps illustrating cross-linked peptides according to nitrogen isotype. In B, E, and F, maps were generated in XiView proteins categorized based on MitoCarta3.0. Data in C, D, and F are mean ± SEM; n = 2 mice. *, P < 0.05; **, P < 0.01, by Kruskal–Wallis ANOVA with Tukey’s multiple comparisons test. See also Fig. S5.