The β(2)-adrenergic receptor (ADRB2) mediates obesity, cardiorespiratory fitness, and insulin resistance. We examined the hypothesis that ADRB2 Arg16Gly-Gln27Glu haplotype is associated with body composition, glucose tolerance, and insulin sensitivity in obese, postmenopausal women. Obese (>35% body fat), postmenopausal (age 45-75 years) women (n = 123) underwent genotyping, dual-energy X-ray absorptiometry, and computed tomography scans, exercise testing (VO(2(max))), 2-h oral glucose tolerance tests (OGTTs), and hyperinsulinemic-euglycemic clamps (80 mU/m(2)/min). Analysis of covariance (ANCOVA) tested for differences among haplotypes, with race, % body fat, and VO(2(max)) as covariates. We found that ADRB2 haplotype was independently associated with % body fat, abdominal fat distribution, VO(2(max)), insulin sensitivity (M/ΔInsulin), and glucose tolerance (ANOVA, P < 0.05 for all). Women homozygous for Gly16-Gln27 haplotype had the highest % body fat (52.7 ± 1.9%), high abdominal fat, low M/ΔInsulin (0.49 ± 0.08 mg/kg/min/pmol/l/10(2)), and impaired glucose tolerance (IGT) during an OGTT (G(120) = 10.2 ± 0.9 mmol/l). Women homozygous for Gly16-Glu27 haplotype also had low M/ΔInsulin (0.51 ± 0.05 mg/kg/min/pmol/l/10(2)) and IGT (G(120) = 8.2 ± 0.7 mmol/l). Subjects with Arg16-Gln27/Gly16-Gln27 haplotype combination had the highest VO(2(max)) (1.84 ± 0.07 l/min) and M/ΔInsulin (0.7 ± 0.04 mg/kg/min/pmol/l/10(2)), and normal glucose tolerance (G(120) = 6.4 ± 0.4 mmol/l), despite being obese. These data show associations of the ADRB2 Arg16Gly-Gln27Glu haplotype with VO(2(max)) and body composition, and an independent association with glucose metabolism, which persists after controlling for body composition and fitness. This suggests that ADRB2 haplotypes may mediate insulin action, glucose tolerance, and potentially risk for type 2 diabetes mellitus (T2DM) in obese, postmenopausal women.