Mirror-image Dependence: Targeting Enantiomeric G-quadruplex DNA by 10 Pairs of Triplex Metallohelices

DNA chirality is the fundamental issue for design of DNAtargeted chiral drugs and DNA-based chiral devices. Natural D-DNA and L-DNA are mirror-image counterparts. However, due to the inherent flexibility and conformation diversity of DNA, it is still not clear how enantiomeric compounds to recognize D-DNA and L-DNA. Are their recognitions following mirror-image dependence? Herein, taken G-quadruplex (G4) DNA as an example which has diverse conformations and distinct biofunctions, 10 pairs of iron triplex metallohelices binding to Dand L-G4 DNA have been evaluated. Screening results clearly show that Δ-enantiomer binding to D-DNA and Λ-enantiomer binding to L-DNA exhibits almost the same stabilization effect and binding affinity. In terms of G4 stabilization effect, iron metallohelices 3 displays the best chiral selectivity on antiparallel G4 DNA, one enantiomer stabilizing G4 DNA while the other even destabilizing G4 DNA. The binding affinity of Δ-3 with DG4 is nearly 70-fold higher than that of Λ-3 with D-G4. Δ-3 binding to D-G4 is following two-step binding and driven by a favourable enthalpy contribution to compensate the associated unfavourable entropy penalty. This work will advance current understanding of DNA chiral recognition and promote DNA-based chiral drug design. Evolution of handedness is a characteristic hallmark of terrestrial life. Chiral recognition of biomolecules is a prerequisite in most biological processes. [1] As the carrier of genetic information, DNA possesses intrinsic chirality. [2] Unraveling the recognition mechanism of chiral species with DNA is an essential determinant for designing DNA-targeted drugs and developing DNA-based devices for chiral applications, including chiral sensing, chiral separation, asymmetric catalysis, and construction of molecular machine and logic devices. [3] L-DNA, as the mirror-image counterpart of natural DNA (DDNA), has identical physical characteristics to D-DNA but with opposite chirality. [4] Similar to D-DNA, L-DNA is polymorphic and can form various secondary structures, such as B-DNA, Z-DNA and G-quadruplex (G4) DNA, etc.[5] But distinctive from D-DNA, L-DNA is nuclease-resistant for its nonnative chirality and thus can be survived in biological environment. [6] L-DNA has been considered to be appealing replacement of D-DNA in nucleic acids-based in vivo diagnostic and therapeutic applications. [7] However, due to the inherent flexibility and conformation diversity of DNA, it is still unclear how enantiomeric compounds to recognize D-DNA and L-DNA. Are their recognitions following mirror-image dependence? To this end, we choose G4 DNA as an example because G4 DNA is one of the most important noncanonical DNA with diverse conformations and distinct biofunctions. [8] Herein, 10 pairs of iron (Fe2+) chiral barrel-like triplex metallohelices (metallohelices 1-10), bearing both small molecular chemical features and helical nanometer size of a zinc-finger-like DNAbinding motif, binding to human telomeric G4 DNA (abbreviated as D-G4) and L-G4 have been screened and systematically compared with their enantioselectivity (Figure 1 and Figure S1). The screening results clearly indicated that Δ-enantiomer binding to D-G4 and Λ-enantiomer binding to L-G4 exhibited almost the same G4 DNA stabilization effect and binding affinity. In terms of G4 stabilization effect, metallohelices 3 displayed the best chiral selectivity on antiparallel G4 DNA, where one enantiomer stabilizing G4 while the other even destabilizing G4. To our knowledge, this is the first example to identify that it is mirror-image dependent for enantiomeric compounds to recognize Dand L-DNA. CD studies verified that L-Tel22 (L-DNA with the same sequence as human telomeric D-Tel22) could form mirror-image structures of both antiparallel and hybrid D-G4 (Figure S2),[9] indicative of the formation of left-handed L-G4 (Figure 1B). [10] Next, screening of 10 pairs of metallohelices binding to Dand L-G4 was performed. [11] Binding constants of Dand L-G4 (antiparallel G4 in Na+ buffer and hybrid G4 in K+ buffer) with metallohelices were measured by UV-visible titration experiments. [12] For comparison, the binding constants were summarized in Figure 2A and 2B. Obviously, for all metallohelices, the binding constants of D-G4/Δ-enantiomers were nearly identical to that of L-G4/Λ-enantiomers. The same trend was found for L-G4/Δ-enantiomers and D-G4/Λenantiomers, no matter antiparallel or hybrid G4 used. Therefore, it can be concluded that the affinity of D/Δ and L/Λ pair is stronger than that of D/Λ and L/Δ pair, supporting the chiral selectivity, where left-handed Λ-enantiomer prefer left-handed LG4, and right-handed Δ-enantiomer like right-handed D-G4. The effects of metallohelices on the thermal stability of Dand L-G4 were next investigated (Figure S3-S6). DNA melting temperature change (ΔTm) caused by metallohelices were shown in Figure 2C and 2D. Δ-enantiomers of metallohelices 1-3 displayed a better stabilization effect on both antiparallel and hybrid D-G4 than metallohelices 4-10. Metallohelices 5-7 [a] C. Zhao, A. Zhao, Prof. J. Ren, and Prof. X. Qu Laboratory of Chemical Biology and State Key Laboratory of Rare Earth Resource Utilization, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun, Jilin 130022, P. R. China E-mail: xqu@ciac.ac. cn [b] H. Song, Prof. P. Scott Department of Chemistry, University of Warwick, Gibbet Hill Road, Coventry CV4 7AL, United Kingdom [c] A. Zhao University of Chinese Academy of Sciences, Beijing 100039, P. R. China [d] Dr. Hisae Tateishi-Karimata, and Prof. Naoki Sugimoto FIBER, Konan University, 7-1-20 Minatojima-minamimachi, Chuo-ku, Kobe 650-0047, Japan Supporting information for this article is given via a link at the end of the document.