Single Molecule Force Spectroscopy of DNA Complexed with Drugs

R. Krautbauer1 T. E. Schrader1, H. Clausen-Schaumann1, L. H. Pope2, S. Allen2, and H. E. Gaub1

1Lehrstuhl für angewandte Physik, Ludwig-Maximilians Universität & CeNS, Amalienst. 54, D-80799 München, Germany, and 2School of Pharmaceutical Sciences, The University of Nottingham, Nottingham NG7 2RD, United Kingdom


Individual double stranded DNA molecules undergo a highly cooperative structural transition if they are stretched with forces of 65-70 pN, leading to an overstretched structure (S-DNA). If the molecules are stretched with higher forces a second structural transition (melting transition) is observed, in which the double strand is mechanically separated, leaving only a single strand attached between AFM-tip and substrate. These well known features were used to characterize the changes in DNA mechanics upon binding of different drugs.

Cisplatin (cis-diamminedichloroplatinum(II)) can introduce intrastrand crosslinks at dGdG, dGdXdG and dAdG sequences, whereas interstrand crosslinking can occur at dGdC-steps. Stretching experiments showed that the cooperativity of the BS-transition is reduced in all DNA molecules with bifunctional cisplatin adducts. In poly(dG)·poly(dC) and poly(dG-dC)·poly(dG-dC) DNA the drug inhibits the high force melting transition, whereas in pure poly(dA-dT)·poly(dA-dT) DNA force-induced strand separation seems still possible. In poly(dA-dC)·poly(dG-dT) DNA a high force transition is present, but no hysteresis between stretching and relaxing the molecules can be observed.

Binding of the intercalator Ethidium Bromide disrupts the cooperativity of the BS-transition in a characteristic manner. No differences in the effects to fragments of lambda phage DNA and poly(dG)·poly(dC) are observed. At higher concentrations of the drug the force induced melting can no longer be observed.

The minor grove binder berenil shows again different effects on the force spectra. In the low force regime the molecules become stiffer and the BS-transition looses cooperativity. At high concentrations the force at the end of the BS-transition is largely increased and mechanical strand separation is inhibited.

As all force spectra were characteristic for each drug, this technique might also become useful to investigate the interaction of other binding agents and drugs with DNA.