The fat‐1 gene of C. elegans encodes a desaturase, which can convert n‐6 to n‐3 fatty acids. A transgenic mouse that constitutively expresses this gene was used as a mammalian model of increased tissue n‐3 fatty acids. The aim of the study was to evaluate the influence of changes in membrane fatty acid composition on the lipid‐protein interaction, specifically on the production of H2O2 and electron transport chain enzyme activities. Liver mitochondria were isolated by differential centrifugation and the activities of the electron transport chain enzymes determined spectrophotometrically, while H2O2 production was determined fluorimetrically. Fatty acid analyses of mitochondrial membranes were undertaken on Percoll‐purified mitochondria, and performed by Lipomics Technology, Inc. The results indicate that the transgenic mice show a decreased H2O2 production at complex I, as well as decreased complex I enzyme activity. Total H2O2 production was also decreased when mitochondria were respiring on succinate or succinate/glutamate/malate. Fatty acid analysis showed an increased n‐3/n‐6 ratio in mitochondrial PC, PE and LyPC lipid classes, with arachidonic acid (n‐6) decreased and eicosapentaenoic acid (n‐3) increased in all phospholipid classes. The results indicate that decreased complex I‐linked ROS production may provide some protection against oxidative stress in membranes with high n‐3 levels.