Induced fit conformational changes of a "reversed amidine" heterocycle: optimized interactions in a DNA minor groove complex - PubMed (original) (raw)

. 2007 May 2;129(17):5688-98.

doi: 10.1021/ja069003n. Epub 2007 Apr 11.

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Induced fit conformational changes of a "reversed amidine" heterocycle: optimized interactions in a DNA minor groove complex

Manoj Munde et al. J Am Chem Soc. 2007.

Abstract

To better understand the molecular basis for recognition of the DNA minor groove by heterocyclic cations, a series of "reversed amidine" substituted heterocycles has been prepared. Amidine derivatives for targeting the minor groove have the amidine carbon linked to a central heterocyclic system, whereas in the reverse orientation, an amidine nitrogen provides the link. The reverse system has a larger dihedral angle as well as a modified spatial relationship with the groove relative to amidines. Because of the large dihedral, the reversed amidines should have reduced binding to DNA relative to similar amidines. Such a reduction is observed in footprinting, circular dichroism (CD), biosensor-surface plasmon resonance (SPR), and isothermal titration calorimetric (ITC) experiments with DB613, which has a central phenyl-furan-phenyl heterocyclic system. The reduction is not seen when a pyrrole (DB884) is substituted for the furan. Analysis of a number of derivatives defines the pyrrole and a terminal phenyl substituent on the reversed amidine groups as critical components in the strong binding of DB884. ITC and SPR comparisons showed that the better binding of DB884 was due to a more favorable binding enthalpy and that it had exceptionally slow dissociation from DNA. Crystallographic analysis of DB884 bound to an AATT site shows that the compound was bound in the minor groove in a 1:1 complex as suggested by CD solution studies. Surprisingly, unlike the amidine derivative, the pyrrole -NH of DB884 formed an H-bond with a central T of the AATT site and this accounts for the enthalpy-driven strong binding. The structural results and molecular modeling studies provide an explanation for the differences in binding affinities for related amidine and reversed amidine analogues.

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Figures

Figure 1

Figure 1

DNA oligomer sequences used in this study and representative diamidines.

Figure 2

Figure 2

DNase I footprinting titration experiments. The 117-bp EcoRI-PVuII restriction fragment from plasmid pBS was 3′-end-labeled at the EcoRI site with [a-32P]dATP in the presence of AMV reverse transcriptase. The products of the DNase I digestion were resolved on an 8% polyacrylamide gel containing 8 M urea. Drug concentrations are at the top of the lanes. Tracks labeled G represent dimethylsulfate piperidine markers specific for guanines. Sequences at the footprinting sites are indicated to the right of the gel.

Figure 3

Figure 3

CD spectra of DB884, DB613 and DB890 with d(GCGAATTCGC)2. (A) In DB884 the ratios of the compound to DNA from the bottom to the top at 375 nm 0, 0.25, 0.5, 0.75, 1.0 and 1.2. (B) In DB613 the ratios of compound to DNA are 0, 0.25, 0.5, 0.75, 1.0 and 1.2. (C) In DB890 the ratios of compound to DNA are 0.5, 0.75, 1.0, 1.25, 1.75, 2.75. The experiments were conducted in cacodylic acid buffer at 25 °C.

Figure 4

Figure 4

SPR sensograms for (A) DB884 with AATT hairpin. The compound concentrations were 0.0, 0.01, 0.02, 0.06, 0.065, 0.07, 0.75, 0.08, 0.09, 0.1 uM from bottom to top. (B) DB613 with AATT hairpin DNA. The compound concentrations were 0.05, 0.1 uM from bottom to top. (C) RU values are plotted against the unbound compound concentration, Cf (flow solution) for DB884 (closed circles) and DB613 (closed squares) binding to AATT DNA hairpin. The data was fitted to one site model using equation 1.

Figure 5

Figure 5

ITC experimental curves at 25 °C for titration of 0.01 mM (A) DB884 and (B) DB613 into 0.001 mM d(GCGAATTCGC)2 duplex. Results were converted to molar heats and plotted against the compound/DNA molar ratio. The line shows the fit to the results and gives best fit ΔH values for binding.

Figure 6

Figure 6

A 2Fo - Fc electron density map with contours drawn at the 1.3σ level, viwed down onto the DB 884 molecule in the DNA minor groove, and showing some of the neighbouring nucleotides and water molecules. Drawn with the Turbo-Frodo program.

Figure 7

Figure 7

Structure of the DB884-DNA complex, showing the solvent-accessible surface coloured according to crystallographic temperature factors (B values), with red indicating the highest values. The bonds of the DB884 molecule are drawn in green, and its solvent-accessible surface is shown with 50% transparency to heighten clarity. Note close contact of the surfaces of the two terminal phenyl groups of the ligand with the minor groove wall surfaces. Figure has been drawn with the Chimera program.

Figure 8

Figure 8

Detailed views of the hydrogen-bond interactions between the DB884 ligand (drawn in mauve) and the DNA. Distances are in Å. Figures have been drawn with the Chimera program. (A) showing the three hydrogen bonds between amidinium nitrogen atom N20 and acceptor atoms O2, O4′ and a phosphate oxygen atom. Also shown is the network of hydrogen bonds bridging the two water molecules W59 and W60 with the outer-facing amidinium nitrogen atom N18 and N2 of Gua18. (B) Showing the hydrogen bond between the central pyrrole ring nitrogen atom of DB884, and the O2 atom of Thy19. (C) Showing the hydrogen bonds between the amidinium nitrogen atom N28 and the O2 atoms of Thy20 and Cyt21.

Figure 9

Figure 9

DFT ab initio calculations at the 631G (p,d) approximation level for DB884 in the free (top) and bound (bottom) form with their respective electrostatic potential maps.

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References

    1. Tidwell RR, Boykin DW. In: Dicationic DNA minor-groove binders as antimicrobial agents. In DNA and RNA Binders: From Small Molecules to Drugs. Demeunynck M, Bailly C, Wilson WD, editors. Vol. 2. WILEY-VCH; Weinheim: 2003. pp. 414–460.
    1. Wilson WD, Nguyen B, Tanious FA, Mathis A, Hall JE, Stephens CE, Boykin DW. Curr Med Chem Anti-Canc Agents. 2005;5:389–408. - PubMed
    1. Wilson WD, Tanious FA, Ding D, Kumar A, Boykin DW, Colson P, Houssier C, Bailly C. J Am Chem Soc. 1998;120:10310–10321.
    1. Laughton CA, Tanious F, Nunn CM, Boykin DW, Wilson WD, Neidle S. Biochemistry. 1996;35:5655–61. - PubMed
    1. Kopka ML, Yoon C, Goodsell D, Pjura P, Dickerson RE. Proc Natl Acad Sci U S A. 1985;82:1376–1380. - PMC - PubMed

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