HgII binds to C-T mismatches with high affinity.

ABSTRACT: Binding reactions of HgII and AgI to pyrimidine-pyrimidine mismatches in duplex DNA were characterized using fluorescent nucleobase analogs, thermal denaturation and 1H NMR. Unlike AgI, HgII exhibited stoichiometric, site-specific binding of C-T mismatches. The on- and off-rates of HgII binding were approximately 10-fold faster to C-T mismatches (kon ? 105 M-1 s-1, koff ? 10-3 s-1) as compared to T-T mismatches (kon ? 104 M-1 s-1, koff ? 10-4 s-1), resulting in very similar equilibrium binding affinities for both types of 'all natural' metallo base pairs (Kd ? 10-150 nM). These results are in contrast to thermal denaturation analyses, where duplexes containing T-T mismatches exhibited much larger increases in thermal stability upon addition of HgII (?Tm = 6-19°C), as compared to those containing C-T mismatches (?Tm = 1-4°C). In addition to revealing the high thermodynamic and kinetic stabilities of C-HgII-T base pairs, our results demonstrate that fluorescent nucleobase analogs enable highly sensitive detection and characterization of metal-mediated base pairs - even in situations where metal binding has little or no impact on the thermal stability of the duplex.

SUBMITTER: Schmidt OP 

PROVIDER: S-EPMC6061796 | biostudies-other | 2018 Jul

REPOSITORIES: biostudies-other