Abstract Background: Loss-of-function mutations in the SGLT1 (sodium/glucose co-transporter-1) gene result in a rare glucose/galactose malabsorption disorder and neonatal death if untreated. In the general population, variants related to intestinal glucose absorption remain uncharacterized. Objectives: The goal of this study was to identify functional SGLT1 gene variants and characterize their clinical consequences. Methods: Whole exome sequencing was performed in the ARIC (Atherosclerosis Risk in Communities) study participants enrolled from 4 U.S. communities. The association of functional, nonsynonymous substitutions in SGLT1 with 2-h oral glucose tolerance test results was determined. Variants related to impaired glucose tolerance were studied, and Mendelian randomization analysis of cardiometabolic outcomes was performed. Results: Among 5,687 European-American subjects (mean age 54 ± 6 years; 47% male), those who carried a haplotype of 3 missense mutations (frequency of 6.7%)—Asn51Ser, Ala411Thr, and His615Gln—had lower 2-h glucose and odds of impaired glucose tolerance than noncarriers (β-coefficient: −8.0; 95% confidence interval [CI]: −12.7 to −3.3; OR: 0.71; 95% CI: 0.59 to 0.86, respectively). The association of the haplotype with oral glucose tolerance test results was consistent in a replication sample of 2,791 African-American subjects (β = −16.3; 95% CI: −36.6 to 4.1; OR: 0.39; 95% CI: 0.17 to 0.91) and an external European-Finnish population sample of 6,784 subjects (β = −3.2; 95% CI: −6.4 to −0.02; OR: 0.81; 95% CI: 0.68 to 0.98). Using a Mendelian randomization approach in the index cohort, the estimated 25-year effect of a reduction of 20 mg/dl in 2-h glucose via SGLT1 inhibition would be reduced prevalent obesity (OR: 0.43; 95% CI: 0.23 to 0.63), incident diabetes (hazard ratio [HR]: 0.58; 95% CI: 0.35 to 0.81), heart failure (HR: 0.53; 95% CI: 0.24 to 0.83), and death (HR: 0.66; 95% CI: 0.42 to 0.90). Conclusions: Functionally damaging missense variants in SGLT1 protect from diet-induced hyperglycemia in multiple populations. Reduced intestinal glucose uptake may protect from long-term cardiometabolic outcomes, providing support for therapies that target SGLT1 function to prevent and treat metabolic conditions.