In type 2 diabetes, glucose phosphorylation, a regulatory step in glucose utilization by skeletal muscle, is impaired. Since glucokinase expression in skeletal muscle of transgenic mice increases glucose phosphorylation, we examined whether such mice counteract the obesity and insulin resistance induced by 12 wk of a high-fat diet. When fed this diet, control mice became obese, whereas transgenic mice remained lean. Furthermore, high-fat fed control mice developed hyperglycemia and hyperinsulinemia (a 3-fold increase), indicating that they were insulin resistant. In contrast, transgenic mice were normoglycemic and showed only a mild increase in insulinemia (1.5-fold). They also showed improved whole body glucose tolerance and insulin sensitivity and increased intramuscular concentrations of glucose 6-phosphate and glycogen. A parallel increase in uncoupling protein 3 mRNA levels in skeletal muscle of glucokinase-expressing transgenic mice was also observed. These results suggest that the rise in glucose phosphorylation by glucokinase expression in skeletal muscle leads to increased glucose utilization and energy expenditure that counteracts weight gain and maintains insulin sensitivity.