Influence of substituent modifications on the binding of 2-amino-1,8-naphthyridines to cytosine opposite an AP site in DNA duplexes: thermodynamic characterization

Abstract

Here, we report on a significant effect of substitutions on the binding affinity of a series of 2-amino-1,8-naphthyridines, i.e., 2-amino-1,8-naphthyridine (AND), 2-amino-7-methyl-1,8-naphthyridine (AMND), 2-amino-5,7-dimethyl-1,8-naphthyridine (ADMND) and 2-amino-5,6,7-trimethyl-1,8-naphthyridine (ATMND), all of which can bind to cytosine opposite an AP site in DNA duplexes. Fluorescence titration experiments show that the binding affinity for cytosine is effectively enhanced by the introduction of methyl groups to the naphthyridine ring, and the 1:1 binding constant (106 M−1) follows in the order of AND (0.30) < AMND (2.7) < ADMND (6.1) < ATMND (19) in solutions containing 110 mM Na+ (pH 7.0, at 20°C). The thermodynamic parameters obtained by isothermal titration calorimetry experiments indicate that the introduction of methyl groups effectively reduces the loss of binding entropy, which is indeed responsible for the increase in the binding affinity. The heat capacity change (ΔCp), as determined from temperature dependence of the binding enthalpy, is found to be significantly different between AND (−161 cal/mol K) and ATMND (−217 cal/mol K). The hydrophobic contribution appears to be a key force to explain the observed effect of substitutions on the binding affinity when the observed binding free energy (ΔGobs) is dissected into its component terms.

Recent Progress in Abasic Site-binding Small Molecules for Detecting Single-base Mutations in DNA

“…Herein, a benzofurazan derivative, 4-(N,Ndimethylaminosulfonyl)-1,2,3-benzoxadiazole (DBD), 62 was utilized as the environmentally sensitive fluorescent dye, which is linked through an alkyl spacer to 2-amino-5,6,7-trimethyl-1,8-naphthyridine (ATMND). 12 The resulting conjugate with an ethylene linker, ATMND-DBD (Figs. 3A and 3B), was used simultaneously with the AP-containing DNA probe as described in the following two steps: (i) a target single-stranded DNA is hybridized with the AP-containing DNA probe to place the AP site opposite a nucleobase to be detected, (ii) the ligand (ATMND moiety) selectively binds to the orphan (target) nucleobase in the duplex through complementary hydrogenbonding, and the DBD moiety becomes located at the surface of the minor groove of the duplex.…”

“…in DNA duplexes, [10][11][12][13][14][15][16][17][18][19][20][21] and have proposed a new strategy of ligand-based fluorescence assay for SNP typing. In contrast to typical DNA-binding ligands capable of targeting double stranded DNAs by intercalation or groove binding, 22,23 it is characteristic of ligands to bind to non-Watson-Crick basepairing sites in DNA duplexes, where the binding is promoted by a pseudo-base pairing along the Watson-Crick edge of intrahelical target nucleobases, and also by stacking with two nucleobases flanking the AP site.…”

“…In contrast to typical DNA-binding ligands capable of targeting double stranded DNAs by intercalation or groove binding, 22,23 it is characteristic of ligands to bind to non-Watson-Crick basepairing sites in DNA duplexes, where the binding is promoted by a pseudo-base pairing along the Watson-Crick edge of intrahelical target nucleobases, and also by stacking with two nucleobases flanking the AP site. Useful affinity and selectivity for target nucleobases opposite the AP site has been indeed obtained by various kinds of heterocyclic planer compounds, including cytosine-selective 2-amino-1,8-naphthyridine derivatives, [10][11][12][13][14] guanine-selective pterin derivatives, [15][16][17] adenineselective 6,7-dimethyllumazine, 18 and thymine-selective 3,5-diaminopyrazine derivatives. [19][20][21] All of these ligands show a complexation-induced fluorescence signaling, and genotype of samples can be clearly distinguished by combining ligands with selectivity for respective target nucleobases.…”

Competitive Binding of Abasic Site-Binding Ligands and Masking Ligands to DNA Duplexes for the Analysis of Single-Base Mutation

“…13,14 In this direction, we have developed a series of abasic site (apyrimidinic or apurinic; AP site)-specific binding ligands with fluorescence signaling. [15][16][17][18][19][20][21][22] These ligands can strongly and selectively bind to the target nucleobase opposite an AP site by pseudo-base pairing through hydrogen bonding. The binding events are further promoted by stacking interactions with the nucleobases flanking the AP site.…”

“…1A) is the promising cytosine (C)-selective candidate for SNPs typing, because of its strong binding affinity (the dissociation constant Kd = 53 nM at 20 C, I = 110 mM, pH 7.0). 15 C-selectivity of ATMND is rationalized by protonation at the N1 position in the naphthyridine ring to produce a hydrogen-bonding surface fully complementary to that of C (Fig. 1B).…”