New Insights into Quinine–DNA Binding Using Raman Spectroscopy and Molecular Dynamics Simulations

Abstract

Quinine’s ability to bind DNA and potentially inhibit transcription and translation has been examined as a mode of action for its antimalarial activity. UV absorption and fluorescence-based studies have lacked the chemical specificity to develop an unambiguous molecular-level picture of the binding interaction. To address this, we use Raman spectroscopy and molecular dynamics (MD) to investigate quinine—DNA interactions. We demonstrate that quinine’s strongest Raman band in the fingerprint region, which derives from a symmetric stretching mode of the quinoline ring, is highly sensitive to the local chemical environment and pH. The frequency shifts observed for this mode in solvents of varying polarity can be explained in terms of the Stark effect using a simple Onsager solvation model, indicating that the vibration reports on the local electrostatic environment. However, specific chemical interactions between the quinoline ring and its environment, such as hydrogen bonding and π-stacking, perturb the frequency of this mode in a more complicated but predictable manner. We use this vibration as a spectroscopic probe to investigate the binding interaction between quinine and DNA. We find that, when the quinoline ring is protonated, quinine weakly intercalates into DNA by forming π-stacking interactions with the base pairs. The Raman spectra indicate that quinine can intercalate into DNA with a ratio reaching up to roughly one molecule per 25 base pairs. Our results are confirmed by MD simulations, which also show that the quinoline ring adopts a t-shaped π-stacking geometry with the DNA base pairs, whereas the quinuclidine head group weakly interacts with the phosphate backbone in the minor groove. We expect that the spectral correlations determined here will enable future studies to probe quinine’s antimalarial activities, such as disrupting hemozoin biocrystallization, which is hypothesized to be, among other things, one of its primary modes of action against Plasmodium parasites.

Quinine copolymer reporters promote efficient intracellular DNA delivery and illuminate a protein-induced unpackaging mechanism

“…The mechanism underlying the exceptional compaction of pDNA by quinine in poly(quinine- co- HEA) was further investigated using Raman spectroscopy. We previously proved that the quinoline ring symmetric stretching mode (referred to as the quinoline ring mode henceforth) quantitatively reports on the local interactions of quinine with its chemical environment ( 26 ). In the case of monomeric quinine, the quinoline ring mode characteristically upshifts in frequency upon deintercalation from DNA due to the loss of π-stacking interactions with nucleobases ( SI Appendix , Table S3 ).…”

“…Quinine Facilitates Enhanced Binding to DNA in Polyplexes. Our previous work shows that monomeric quinine interacts with DNA electrostatically via the quinuclidine moiety, as well as through intercalation via the quinoline ring (26). We hypothesized that the quinine copolymers would efficiently bind and compact pDNA and form self-assembled polyplexes in solution via both binding modes, allowing for protection against degradation and efficient cellular delivery.…”

“…In addition to promoting endosomal escape ( 21 – 23 ), chloroquine has been shown to facilitate unpackaging of pDNA by competitive binding ( 24 , 25 ). Like chloroquine, the structurally similar Cinchona alkaloid, quinine, can bind DNA through both electrostatic interactions with the phosphate backbone and intercalation through π-stacking interactions with nucleobases ( 26 , 27 ). Therefore, incorporating quinine into a polymer would enable robust DNA binding and polyplex stabilization via multiple binding mechanisms, which minimizes the overall charge density of the polymer necessary to compact DNA.…”

“…A major advantage of Raman spectroscopy is that it monitors vibrations that are intrinsically sensitive to the structure and interactions of molecules ( 36 – 41 ). We recently showed, for example, that the Raman bands of quinine show characteristic frequency shifts that report on its electrostatic, hydrogen bonding, and π-stacking interactions, which can be used to monitor its intercalation with DNA ( 26 ). Despite this, the rich chemical information encoded in Raman spectra has surprisingly not been exploited in imaging applications to probe how macromolecules dynamically interact with their local physiochemical environment in situ.…”

Stimulated Raman imaging below the diffraction limit with a MHz laser

“…[35,36] These tags have a drastically lower probability of perturbing the system of interest than fluorophores, which are typically on the 1-nm length scale for organic dye molecules and the 10-nm length scale for fluorescent proteins. Additionally, Raman spectra contain information about the surrounding chemical environment [37][38][39][40][41][42] and offer improved multiplexing capabilities due the peaks' narrow bandwidth when compared to fluorescence peaks. [43] However, because Raman scattering is a weak process, standard Raman microscopy has limited use as a biological imaging technique.…”