Nanoporous gold (NPG) prepared by de-alloying Al2Au exhibits correlation between the high catalytic reactivity towards CO oxidation and the density of twinning defects in the fcc lattice of NPG. It was also discovered that on the internal surface of NPG, quite common twinning defects can create close-packed rows of six-coordinated catalytically active Au atoms denoted as W-chains. In this work, using density functional theory methods, we investigate energy conditions for formation, thermal stability, and chemical reactivity of these active sites. The possibility of dioxygen chemisorption on various surface sites is studied in detail. A contribution from the dispersion interactions is also considered. The calculated surface density of the active six-coordinated atoms in NPG comparable with that of supported gold nanoparticle catalysts, exothermic chemisorption of dioxygen, and the energy profiles of reaction pathways for CO oxidation indicate that the six-coordinated sites created by twinning can significantly contribute to the catalytic activity of NPG.