Pyruvate kinase is a key glycolytic enzyme. Isoforms that are expressed in the red cell, liver, pancreatic beta-cells, small intestine, and proximal renal tubule are encoded by the 12 exons of the PKLR gene, which maps to chromosome 1q23. We hypothesized that common variants of the PKLR gene could account for the linkage of diabetes to this region. We screened the promoter regions, exons and surrounding introns, and the 3' untranslated region for mutations. We identified five single-nucleotide polymorphisms (SNPs), and only one (V506I, exon 11) altered the coding sequence. We tested the five SNPs, a poly-T insertion-deletion polymorphism, and an ATT triplet repeat in 131 unrelated diabetic patients and 118 nondiabetic control subjects. The V506I variant was rare and not associated with type 2 diabetes. The four SNPs and the insertion-deletion polymorphism were associated with diabetes, with a 10% difference between individuals with diabetes and nondiabetic individuals (P = 0.001-0.011, relative risk for minor allele 1.85). The same trend was found for the ATT repeat (P = 0.029). Common variants in the PKLR are associated with increased risk of type 2 diabetes, but because of strong linkage disequilibrium between variants, the actual susceptibility allele may be in a different gene.