Monitoring DNA-Ligand Interactions in Living Human Cells Using NMR Spectroscopy
| Authors | |
|---|---|
| Year of publication | 2019 |
| Type | Article in Periodical |
| Magazine / Source | Journal of the American Chemical Society |
| MU Faculty or unit | |
| Citation | |
| web | https://pubs.acs.org/doi/pdf/10.1021/jacs.9b03031 |
| Doi | http://dx.doi.org/10.1021/jacs.9b03031 |
| Keywords | MINOR-GROOVE; NUCLEIC-ACID; DRUG DESIGN; DISCOVERY; BINDERS |
| Description | Studies on DNA-ligand interactions in the cellular environment are problematic due to the lack of suitable biophysical tools. To address this need, we developed an in-cell NMR-based approach for monitoring DNA-ligand interactions inside the nuclei of living human cells. Our method relies on the acquisition of NMR data from cells electroporated with preformed DNA-ligand complexes. The impact of the intracellular environment on the integrity of the complexes is assessed based on in-cell NMR signals from unbound and ligand-bound forms of a given DNA target. This technique was tested on complexes of two model DNA fragments and four ligands, namely, a representative DNA minor-groove binder (netropsin) and ligands binding DNA base-pairing defects (naphthalenophanes). In the latter case, we demonstrate that two of the three in vitro-validated ligands retain their ability to form stable interactions with their model target DNA in cellulo, whereas the third one loses this ability due to off-target interactions with genomic DNA and cellular metabolites. Collectively, our data suggest that direct evaluation of the behavior of drug-like molecules in the intracellular environment provides important insights into the development of DNA-binding ligands with desirable biological activity and minimal side effects resulting from off-target binding. |
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