Figure 3.

Different factors affecting observed pK_a. A) ¹P NMR-based pH titration over a wide pH range (4.0-12.0) of a 19 mer dsDNA containing an A⁺•C wobble base pair. The plot shows five regions: acid denaturation (negative slope between pH ∼3.8-4.5), protonated folded state (flat slope between pH ∼4.5-6.0), ionization of the wobble base pair (positive slope between pH∼6.0-8.0), deprotonated folded state (flat slope between pH ∼8.0-10.0), and alkaline dentaturation (positive slope between pH ∼10.0-12.0). Adapted with permission from Siegfried et al. 2010. Copyright (2010) American Chemical Society. B) ³¹P NMR-based pH titration of DNA with interference from alkaline denaturation. The data above pH 9.4 were not included in the fit to Eq. (3) to obtain pK_a. Adapted with permission from Wilcox et al. 2013. Copyright (2013) American Chemical Society. C) The pH-dependent cleavage of RNA under constant Na⁺ concentration. The different lines represent the number of independent ionizations affecting the observed pK_a. The solid line represents 19 independent ionization events each with a microscopic pK_a∼9.4, which gives rise to an apparent pK_a of ∼7.6. Adapted with permission from Knitt et al. 1996. Copyright (1996) American Chemical Society. D) ³¹P NMR-based pH titration of DNA with no interference from unfolding. The observed pK_a is close to neutrality and is easily determined with the ideal complete baselines. Adapted with permission from Wilcox et al. 2013. Copyright (2013) American Chemical Society.