We have previously reported a common variation in the liver promoter of the human glucokinase, which is regulated by insulin, in the patients with non-insulin-dependent diabetes mellitus (NIDDM). The variation occurred within a 10-bp region completely conserved between human and rat. Its basic motif was almost identical to the insulin regulatory element of the phosphoenolpyruvate carboxykinase gene. In vitro transfection experiment showed that the G-to-A variation causes a 58% reduction in the promoter activity. After oral glucose challenge, the homozygous A/A subjects had the highest stimulated insulin levels at 60 and 90 minutes and the highest insulin area under the curve as compared to the subjects with other genotypes, which suggested the homozygous A/A subjects were more insulin resistant. As insulin resistance is a risk factor of NIDDM, we concluded that this promoter variation is a risk factor for NIDDM.