Abstract: Hemoglobin（Hb） plays a key role in oxygen transport in animals. In this study, the members of Hb gene family, the structure of gene cluster and the protein structure of Pyrgilauda ruficollis were studied by genomics, molecular evolution, molecular dynamics and computational biology. The purpose was to explain the mechanism of Hb adaptation to high altitude hypoxia environment in P. ruficollis. The results showed that α（5′-α^E, α^D, α^A-3′） and β（5′-ρ, β^H, β^A, ε-3′）gene clusters of P. ruficollis Hb retained their distribution patterns in the ancestors of birds. Hb α^A had seven mutation sites of potential positive selection, among which 5 sites（p.Pro5Ser、 p.Ala6Ser、 p.Ala27Gly、 p.Asp28Glu、 p.Ala35Thr） mutated from hydrophobic amino acids into hydrophilic amino acids, which was the main reason for hydrophilicity increase of α^A subunit. Mutation of p.Pro45Ala increased the distance between C and F helices, which caused the increase of active center volume of α^A subunit（from 0.683 nm~3 of Passer domesticus to 0.711 nm~3 of Pyrgilauda ruficollis）. The increase of hydrophilicity and active center volume were beneficial to the accumulation of（α^Aβ^A）₂ subtype of Hb and gas exchange in red blood cells. Two mutations including p.Ala27Gly and p.Ala35Thr reduced the number of hydrogen bonds between the dimers of the（α^Aβ^A）₂ subtype of Hb in Pyrgilauda ruficollis, which made the transition of Hb from T state（tense state） to R state（relaxed state） easier. It is speculated that the changes of physical and chemical properties and conformation of Hb caused by the above mutations could improve aerobic affinity of the corresponding subtypes, which may be the main mechanism of Hb adaptation to high altitude hypoxia environment in Pyrgilauda ruficollis.