Theoretical estimations at both the B3LYP/6-3l+G(d)//B3LYP/6-31+G(d) and B3LYP/aug-cc-pVTZ// B3LYP/6-1+G(d) levels for the 298 K gas-phase acidities (GPA) of all the possible deprotonation sites of the most stable tautomers of adenine and guanine are reported in terms of the enthalpy change of deprotonation. The gas-phase acidities of the nucleobases estimated by direct calculations and derived from isodesmic reactions are very similar. At the B3LYP/aug-cc-pVTZ//B3LYP/6-31+G(d) level of theory, for the most stable adenine tautomer, 9H-adenine, the GPAs of N9, N4-H-a, N4-H-b, C8, and C2 sites are 337.8 +/- 2.1, 355.6 +/- 2.1, 355.3 +/- 2.1, 375.7 +/- 0.5, and 401.2 +/- 0.5 kcal/mol, respectively. For the most stable guanine tautomer, 7H-keto-guanine, the GPAs of N3, N7, N2-H-a, N2-H-b, and C8 sites are 336.4 +/- 2.2, 338.0 +/- 2.1, 347.7 +/- 2.1, 341.9 +/- 2.1, and 376.0 +/- 0.5 kcal/mol, respectively. For the second most stable tautomer, 9H-ketoguanine, the GPAs of N3, N9, N2-H-a, N2-H-b, and C8 are 338.6 +/- 2.2, 337.4 +/- 2.1, 343.8 +/- 2.1, 338.4 +/- 2.1, and 379.9 +/- 0.5 kcal/mol, respectively. The GPAs of N9, hydroxyl group, N2-H-a, N2-H-b, and C8 of 9H-cis-enol guanine are 338.7 +/- 2.1, 340.2 +/- 2.1, 358.0 +/- 2.1, 357.7 +/- 2.1, and 378.3 +/- 0.5 kcal/mol, respectively. The corresponding GPAs of 9H-trans-enol guanine are 336.5 +/- 2.1, 339.8 +/- 2.1, 359.1 +/- 2.1, 359.5 +/- 2.1, and 374.6 +/- 0.5 kcal/mol, respectively. The errors given above are based on the experimental uncertainty in the reference values used in calculations involving isodesmic reactions. In general, N9 is the most acidic site for 9H-adenine, followed by the amino group, while C2 is the least acidic site. For both 7H-keto- and 9H-keto-guanine, N3 and N7 (or N9) are the most acidic sites, followed by the amino group. For enol guanine tautomers, N9 is the most acidic site, followed by the hydroxyl group and the amino group. For all guanine tautomers mentioned above, C8 is the least acidic site. Differing from the situation of 9H-adenine and 9H-enol guanine, the acidities of the two amino group hydrogen atoms are significantly different in 7H-keto- and 9H-keto-guanine.