Extended Data Fig. 2.

(a) Structural insight into CX6B1 R20 forming salt-bridges at NDUA4-CX6B1 interface. R20 side chain (green) forms a salt bridge with CX6B1 D16 (green) and NDUA4 D60 (black), which pinpoints an interface necessary for the stability of CIV. Side-chains of R20, D16, and NDUA4 D60 (partially resolved) are depicted in stick representation. Cross-linked lysine sidechains are shown as yellow (CX6B1) or orange (NDUA4) spheres.

(b) Cytochrome C oxidase enzymatic activity assay in tissue homogenates from TAC and Sham hearts, normalized to Citrate Synthase activity. N=4 animals, AVG+/−SEM, *denotes p<0.05 by unpaired, two-tailed Student’s t-test.

(c) Representative Blue Native-PAGE (BN-PAGE) analysis of mitochondria isolated from Sham and TAC groups. Coomassie stain (left) for total protein loading, In-gel CIV activity stain (middle), with CI activity stain overlay (right). Gels were run in duplicates.

(d) Representative BN-PAGE immunoblot of NDUA4 containing SCs from digitonin solubilized isolated mitochondria. Blots were run in duplicates.

(e) Representative BN-PAGE immunoblot of NDUA4 containing SCs from DDM solubilized isolated mitochondria. Blots were run in duplicates.