Rationale: Superoxide dismutase 3 (SOD3) inhibits oxidative fragmentation of lung matrix components collagen I, hyaluronan, and heparan sulfate. Inherited change in SOD3 expression or function could affect lung matrix homeostasis and influence pulmonary function. Objectives and Methods: Resequencing of 182 individuals identified two novel polymorphisms E1 (rs8192287) and I1 (rs8192288) in a conserved region of the SOD3 gene of potential relationship to lung function. We next genotyped 9093 individuals from the Copenhagen City Heart Study for the polymorphisms and recorded spirometry, and admissions and deaths due to COPD during 26 years follow-up. Finally, we validated our findings in a cross-sectional analysis of 35,635 individuals from the Copenhagen General Population Study. Measurements and Main Results: Genotyping the Copenhagen City Heart Study identified 35 E1/I1 homozygotes, 1,050 heterozygotes, and 8,008 non-carriers (Hardy-Weinberg equilibrium:P=0.93). Using quadruple lung function measurements, we found that E1/I1 homozygotes had 7% lower forced vital capacity (FVC)% predicted (P=0.006) and 4% lower forced expiratory volume in first second (FEV₁) %predicted (P=0.12) compared with noncarriers. In the Copenhagen General Population Study, E1/I1 homozygotes also had lower FVC % predicted than noncarriers (P=0.03), confirming an association between E1/I1 genotype and reduced lung function. E1/I1 homozygotes had adjusted hazard ratios for COPD hospitalization and COPD mortality of 2.5 (1.0-5.9) and 3.7 (0.9-15), respectively; the results were independent from influence from the R213G allele of the SOD3 gene. Conclusions: We identified two novel polymorphisms in a conserved region of the SOD3 gene and show that individuals that are homozygous for these polymorphisms have reduced FVC % predicted in two large population-based studies.