<i>Background/Aim:</i> Changes in dietary macronutrient content are involved in altered energy metabolism and obesity development. However, there are controversial views about the obesogenic effects of high-sucrose (HS) diets, which usually lead to obesity and insulin resistance but are sometimes associated with reduced weight gain. The aim of this study was to investigate the effect of consumption of a pair-fed HS diet on weight gain and energy homeostasis in rats, as well as to assess the effects on expression of the NADH dehydrogenase (ubiquinone) 1β subcomplex 6 (NDUFB6) gene in adipose tissue. <i>Methods/Results:</i> Although both dietary groups, i.e. HS and control, were pair-fed (isoenergetic feeding), the HS diet increased adiposity and decreased plasma total and high-density lipoprotein cholesterol levels. While no significant differences were found with regard to serum glucose, insulin, adiponectin, free fatty acids and liver malondialdehyde, a slight increase in serum and liver triglycerides was observed. Interestingly, the gene expression of NDUFB6, an inner mitochondrial membrane protein involved in mitochondrial electron transport, was reduced in epididymal adipose tissue when compared to the control-fed group. <i>Conclusion:</i> These results suggest, apparently for the first time, that high-sugar diets appear to induce mitochondrial dysfunction in adipose tissue, which may be related to greater weight gain and metabolic impairment.