Mitochondria are essential organelles to efficiently produce ATP by ATP-synthase, which uses a proton-gradient generated by respiratory chain complexes. We previously demonstrated that COX7RP/COX7A2L/SCAF1 is a key molecule that promotes respiratory supercomplex assembly and regulates energy generation. The contribution of COX7RP to metabolic homeostasis, however, remains to be clarified. In the present study, we showed a metabolic phenotype of Cox7rp knockout (Cox7rpKO) mice, which exhibit lower blood glucose levels after insulin or pyruvate injection. Notably, ATP synthesis rate was reduced in Cox7rpKO mice liver, in accordance with decreased percentages of complex III subunit RISP and complex IV subunit COX1 involved in I + III + IV supercomplex fraction. The present findings suggest that COX7RP-mediated mitochondrial respiration plays crucial roles in the regulation of glucose homeostasis and its impairment will lead to the pathophysiology of metabolic states.