We here show that the pK(a) (error limit: 0.01 to 0.03 pK(a) unit) of a nucleobase in a nucleotide can be modulated by the chemical nature of the 2'-substituent at the sugar moiety. This has been evidenced by the measurement of nucleobase pK(a) in 47 different model nucleoside 3',5'-bis- and 3'-mono-ethylphosphates. The fact that the electronic character of each of the 2'-substituents ( Fig. 1) alters the chemical shift of the H2' sugar proton, and also alters the pKa of the nucleobase in the nucleotides has been evidenced by a correlation plot of pK(a) of N3 of pyrimidine (T/C/U) or pK(a) of N7 of 9-guaninyl with the corresponding delta H2' chemical shifts at the neutral pH, which shows linear correlation with high Pearson's correlation coefficients ( R = 0.85 - 0.97). That this modulation of the pK(a) of the nucleobase by a 2'-substituent is a through-bond as well as through-space effect has been proven by ab initio determined pK(a) estimation. Interestingly, experimental pK(a)s of nucleobases from NMR titration and the calculated pK(a)s (by ab initio calculations utilizing closed shell HF6-31G** basis set) are linearly correlated with R = 0.98. It has also been observed that the difference of ground and protonated/de-protonated HOMO orbital energies (Delta HOMO, a. u.) for the nucleobases (A/ G/ C/ T/ U) are well correlated with their pK(a)s in different 2'-substituted 3', 5'-bis-ethylphosphate analogs suggesting that only the orbital energy of HOMO can be successfully used to predict the modulation of the chemical reactivity of the nucleobase by the 2'-substituent. It has also been demonstrated that pKa values of nucleobases in 3',5'-bis-ethylphosphates ( Table 1) are well correlated with the change in dipole moment for the respective nucleobases after protonation or de-protonation. This work thus unambiguously shows that alteration of the thermodynamic stability (T-m) of the donor - acceptor complexes [ref. 20], as found with various 2'-modified duplexes in the antisense, siRNA or in triplexes by many workers in the field, is a result of alteration of the pseudoaromatic character of the nucleobases engineered by alteration of the chemical nature of the 2'-substitution.