Monitoring base specific dynamics during melting of DNA-ligand complexes using temperature-jump time-resolved infrared spectroscopy

Fritzsch, Robby and Greetham, Gregory M. and Clark, Ian P. and Minnes, Lucy and Towrie, Michael and Parker, Anthony W. and Hunt, Neil T. (2019) Monitoring base specific dynamics during melting of DNA-ligand complexes using temperature-jump time-resolved infrared spectroscopy. Journal of Physical Chemistry B. ISSN 1520-6106

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    Abstract

    Ultrafast time resolved infrared spectroscopy employing nanosecond temperature-jump initiation has been used to study the melting of double stranded (ds)DNA oligomers in the presence and absence of minor groove-binding ligand Hoechst 33258. Ligand binding to ds(5'-GCAAATTTCC-3'), which binds Hoechst 33258 in the central A-tract region with nanomolar affinity, causes a dramatic increase in the timescales for strand melting from 30 µs to ~250 µs. Ligand binding also suppresses pre-melting disruption of the dsDNA structure, which takes place on 100 ns timescales and includes end-fraying. In contrast, ligand binding to the ds(5'-GCATATATCC-3') sequence, which exhibits an order of magnitude lower affinity for Hoechst 33258 than the A-tract motif, leads to only a factor of five increase in melting timescales and reduced suppression of pre-melting sequence perturbation and end fraying. These results demonstrate a dynamic impact of the minor groove ligand on the dsDNA structure that correlates with binding strength and thermodynamic stabilisation of the duplex. Moreover, the ability of the ligand to influence base pairs distant from the binding site has potential implications for allosteric communication mechanisms in dsDNA.