Figure 2. Respiratory Activities.

(A) SMPs derived from KO liver had no CI activity, whereas CIII and CIV activities were not significantly different. CII activity was significantly higher. Data were normalized to citrate synthase activity (Δ Abs/min of respiratory complex activity : Δ Abs/min citrate synthase activity); hence they have no units. Assays in triplicate, n =12 for CI, CII, CIV; n =4 CIII. *, p =0.003; **, p =0.0001, Mann-Whitney test. Similar results were obtained with SMPs from brain and MEFs. Values given as mean ± s.e.m.

(B) Rates of O₂ consumption by liver CI from WT and KO mice (P35) were measured. Arrows indicate addition of: (1) tissue, (2) ADP, (3) malate/pyruvate, and (4) rotenone. There was less O₂ consumption by KO tissue in the presence of malate/pyruvate (CI) relative to WT, and it stopped with addition of rotenone.

(C) The ratio O₂ consumption (oxygen/mg tissue/min) by liver from KO versus WT mice. Complex I activity of KO tissue is less than half that of WT, whereas CII and CIV activities do not significantly differ, n =5. Values given as mean ± s.e.m. *, p =0.0001, Student’s two-tailed t-test. Similar results were obtained from brain and muscle.

(D) Blue Native Gel Electrophoresis (BNGE) of CI from brain and liver mitochondria. Proteins representing the three major CI subcomplexes were examined by Western blot; in each case, KO mitochondria had less intact complex I.

(E) SDS PAGE of proteins representing the three major subcomplexes of respiratory complex I. Equivalent amounts of mitochondrial protein were loaded, MnSOD was used as internal standard. The samples were the same as used for the BNGE experiment (Fig. 2D).