Soeg K.

Kim, 1
Jian-Sheng Sun, 2
Therese Garestier, 2
Claude Helene, 2
C. H. Nguyen, 3
E. Bisagni, 3
Alison Rodger, 4
Bengt Norden 5
1

Department of Chemistry,
College of Sciences,
Yeungnam University,
Kyoungsan City, Kyoung-buk,
712-749,
Republic of Korea
2

Laboratoire de Biophysique,
Museum National dHistoire
Naturelle,
INSERM U 201, CNRS,
UA 481, 43 rue Cuvier,
75231 Paris Cedex 05, France
3

Laboratoire de Synthese
Organique,
Institut CurieRecherche,
UMR 176 CNRS,
Batiment 110,
91405 Orsay, France

Binding Geometries of Triple

Helix Selective
benzopyrido[4,3-b]indole
Ligands Complexed with
Double- and Triple-Helical
Polynucleotides

Department of Chemistry,
University of Warwick,
Coventry, CV4 7AL, England
5

Department of Physical
Chemistry,
Chalmers University of
Technology,
S 412 96 Gothenburg,
Sweden
Received 13 June 1996;
accepted 2 December 1996

Abstract: The binding modes of three benzopyrido[4,3-b]indole derivatives (and one benzo[ f]pyrido[4-3b]quinoxaline derivative) with respect to double helical poly(dA)rpoly(dT) and
poly[d(A-T)]2 and triple-helical poly(dA)r2poly(dT) have been investigated using linear
dichroism (LD) and CD: (I) 3-methoxy-11-amino-BePI where BePI {7H-8-methyl-benzo[e]pyrido[4,3-b]indole}, (II) 3-methoxy-11-[(3 *-amino)propylamino]-BePI, (III) 3-methoxy-7-[(3 *-diethylamino)propylamino]BgPI where BgPI {benzo[g]pyrido[4,3-b]indole}, and (IV) 3-methoxy-11-[(3 *-amino)propylamino]B f PQ where B f PQ {benzo[ f]pyrido[4-3b]quinoxaline}. The magnitudes of the reduced LD of the electronic transitions
of the polynucleotide bases and of the bound ligands are generally very similar, suggesting an
orientation of the plane of the ligands fused-ring systems preferentially perpendicular to the
helix axis. The LD results suggest that all of the ligands are intercalated for all three polynucleotides. The induced CD spectrum of the BePI chromophore in the (II-BePI)-poly[d(A-T)]2
complex is almost a mirror image of that for the (I-BePI)-poly(dA)rpoly(dT) and (I-BePI)poly(dA)r2poly(dT) complexes, suggesting an antisymmetric orientation of the BePI moiety

Correspondence to: Bengt Norden

q 1997 John Wiley & Sons, Inc.

CCC 0006-3525/97/010101-11

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Kim et al.
upon intercalation in poly[d(A-T)]2 compared to the other polynucleotides. The induced CD
of I-BePI bound to poly(dA)r2poly(dT) suggests a geometry that is intermediate between that
of its other two complexes. The concluded intercalative binding as well as the conformational
variations between the different BePI complexes are of interest in relation to the fact that BePI
derivatives are triplex stabilizers. q 1997 John Wiley & Sons, Inc. Biopoly 42: 101111,
1997
Keywords: triple helix stabilization; intercalation; DNA ligands; benzo[4,3-b]indole; polynucleotides; linear dichroism; CD

INTRODUCTION
The triple-helical structure of nucleic acids was first
documented some 40 years ago.1,2 Recent evidence
on the therapeutic potential of base pair specific
triplex formation 3,4 and potential biological roles of
intramolecular triple helices 5 have caused an explosive interest in studies of triple helical nucleic acids.
Therapeutic use of triplex formation for control of
gene expression may involve inhibition of transcription 6 10 or DNA replication 11,12 or prevention of cellular proteins from binding to their designated target
DNA.13 15 A third strand, for example equipped
with [Fe(II) EDTA] or some ellipticine derivative,
may be used as a site specific endonuclease.16,17 The
biological significance of triplex structures is suggested by the observation, in vivo, of intramolecular
triplex (H-DNA) formation within supercoiled plasmids.18 20
In general, a DNA triple helix is thermodynamically less stable than when it is dissociated into a
double helix and a single-stranded piece of DNA.
In order to enhance the affinity of the third strand for
the duplex under physiological conditions, various
approaches have been taken. These include covalent
linkage to DNA of binding agents, such as intercalators 21,22 and photoinducible cross-linking reagents, 23,24 or addition of triplex-selective ligands
such as benzopyridoindoles that stabilize the triplex
structure.25 28 It is of interest to know where on the
triplex DNA such agents bind. In this paper we use
CD and linear dichroism (LD) to probe the binding
of three benzopyridoindoles molecules and one
benzopyrido quinoxaline molecule to duplex and
triplex DNAs.
In principle, four classes of binding modes in Bform polynucleotide complexes may be discriminatednamely, binding in the minor or major
groove, intercalation, and association on the surface
of the phosphate spine. In addition, there are interactions in which the conformation of DNA is denatured or in other ways severely distorted. Numerous
intercalators and spectroscopic features of their
complexes with DNA have been studied over the
years; most intercalators are polycyclic aromatic
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heterocycles. When these molecules are fully intercalated between adjacent base pairs of a polynucleotide, the latter is expected to become unwound,
elongated, and generally somewhat stiffened. This
would lead to an increase in the magnitude of the
flow oriented LD of the DNA around 260 nm (at
which wavelength the p r p* transitions of the
nucleobases absorb light).29 In an idealized model,
the planes of the intercalated polycyclic aromatic
chromophore and the nucleobases are parallel and
approximately perpendicular to the helix axis. For
comparing the DNA and ligand LDs, the reduced
LD (LD r , the LD divided by normal absorption),
is usually the convenient measure of transition moment orientation. Parallel transition moments have
the same sign and magnitude LD r signals in their
respective absorption regions.29 In practice, the LD r
of an intercalated ligand is often greater (more negative) than the average DNA LD r as the DNA is
stiffer in the region of the intercalated ligand.
The CD spectrum of a ligand, induced by the
interaction of its transition moments with those of
the surrounding chirally arranged polynucleotide
bases, also provides information about the mode of
binding. The induced CD of an intercalated ligand
depends on the orientation of the ligand in the intercalation site, on its displacement from the helix axis,
and also on the identity (including sense) of the
nucleotide step.30,31
One representative minor groove binding ligand
is 4 *,6-diamidino-2-phenylindole (DAPI). Both the
crystal structure of DAPI complexed with the dodecamer, [d(CGCGAATTCGCG)2 ], and spectroscopic studies of its complexes with duplex DNA,
homopolynucleotides, and mixed sequence oligonucleotides, show that DAPI preferentially binds to
the minor groove of AT-rich sequences.29 In the
crystal structure it is inserted edgewise into the docecamer groove in the central AATT stretch. The
LD r of the DAPI-poly[d(A-T)]2 complex is positive in the 300420 nm region, where DAPI is
known to have two electronic transitions polarized
near the long axis of the molecule.29 The long axis
of DAPI is calculated from the LD r to be about
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FIGURE 1 Molecular structures of (a) 3-methoxy-11amino-BePI where BePI {7H-8-methyl-benzo[e]pyrido[4,3-b]indole}, (I); (b) 3-methoxy-11-[(3 *-amino)propylamino]-BePI, (II); (c) 3-methoxy-7-[(3 *-diethylamino)propylamino]BgPI where BgPI {benzo[g]pyrido[4,3-b]indole}, (III); and (d) 3-methoxy-11-[(3 *amino)propylamino]B f PQ where B f PQ {benzo[ f]pyrido[4-3b]quinoxaline} (IV).

457 relative to the polynucleotide helix axis, which

agrees perfectly with the orientation expected for a
minor groove binder given the pitch of the undistorted minor groove. Its short axis lies parallel to the
DNA bases and any short axis polarized transitions
would have a negative LD.
Compared to the minor groove, the major groove
is wide and shallow. There are few known examples
of major groove binding chromophoric ligands and
it is difficult to define any typical spectroscopic
properties of such complexes. The LD r of a major
groove binding ligand could be expected to differ
from that of minor groove bound complexes in that
the size of the major groove allows other orientations than the one along the groove.
The large fused-aromatic ring systems of the
benzopyridoindoles give these compounds strongly
hydrophobic properties and, for example, 3-methoxy-7H-8-methyl-11-[(3 *-amino)propylamino]benzo[e]pyrido [4,3-b]indole [3-methoxy-11-(3 *amino)propylamino-BePI (BePI {7H-8-methylbenzo[e]pyrido[4,3-b]indole}), shown in Figure
1, has been reported to occur as multimers in aqueous solution.28 In conjunction with the large size of
the molecule enabling it to potentially cover a large
fraction of a base triplet, this property might provide
a driving force towards intercalative binding. BePI
itself has been found to unwind negative supercoils
in the pBR322 plasmid and, upon binding to both
double and triple helices, increases their relative
contour lengths.25 27 Energy transfer from poly/

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103

nucleotides bases to the bound BePI molecules have

been observed and the fluorescence of bound BePI
is protected from an acrylamide quencher.26,27 These
results are consistent with BePI intercalating between the bases of both double- and triple-helical
polynucleotides.
In this work, we have studied (I) 3-methoxy11-amino-BePI, (II) 3-methoxy-11- [ (3*-amino)
p r op y l a m i n o ]-B e P I, and (III) 3-m e t h o x y-7[ (3 *-diethylamino)propylamino ]BgPI where
BgPI { benzo [ g]pyrido[4,3-b]indole}] (Figure 1) complexed with double-helical poly(dA)r
poly(dT) and poly[d(A-T)]2 and triple-helical poly(dA)r2poly(dT) using LD and CD. Some results
for a benzo[ f]pyrido[4,3-b]quinoxaline (B f PQ)
derivative (IV in Figure 1) are also included for
purposes of comparison. This combination of techniques has been found useful in providing information about binding geometries and nucleo-base conformations in nucleic acid complexes with ligands
carrying well-characterized anisotropic chromophores.29,32 35 These ligands are related but differ in
some of the features expected to be important for
their binding to DNA. Compound (I) has a single
positive charge on the ring nitrogen while (II) has
a second positive charge on the substituent at position 11. Further, the BePI ring has to adopt a skewed
conformation to avoid clashes between the proton
at position 1 and that of the NH substituent at position 11, whereas the BgPI and B f PQ derivatives
are planar molecules.32 35

MATERIALS AND METHODS

Chemicals
Polynucleotides. Poly(dA), poly(dT), poly(dA)r
poly(dT)and poly[d(A-T)]2 , purchased from Pharmacia, were dissolved in a buffer containing 100 mM
NaCl, 5 mM cacodylate, and 1 mM EDTA, pH 7.0, and
dialyzed several times against 5 mM cacodylate buffer,
pH 6.0. Poly(dA)r2poly(dT) was prepared by simmering a solution containing a 1:2 molar mixture of
poly(dA) and poly(dT) in presence of 2 mM MgCl2 at
907C for 30 min followed by overnight annealing at room
temperature. Formation of the triplex was verified by its
characteristic CD spectrum and melting profile. The concentrations of the polynucleotides were measured spectrophotometrically using molar extinction coefficients of
8600M 01 cm01 (at 257 nm), 8520M 01 cm01 (at 264 nm),
6600M 01 cm01 (at 262 nm), and 6000M 01 cm01 (at 260
nm), respectively, for poly(dA), poly(dT), poly[d(A-T)]2
and poly(dA)rpoly(dT).
Ligands. The ligands were synthesized as reported by
Nguyen et al.32,33 A highly concentrated BePI solution
was prepared in 5 mM cacodylate buffer at pH 6.0 and
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the small aliquots (1020 mL) of this solution were injected into 3 mL of polynucleotide solution (50 mM in
adenine base) to give final ligand concentrations. The
ligand to base pair (for duplex) or base triplet (for triplex) ratio was approximately 0.2. The ligand concentrations were determined spectrophotometrically using the
extinction coefficient of 5500M 01 cm01 at 361 nm for
the BePI chromophore, 6100M 01 cm01 at 361 nm for
BgPI, 28 and 5800M 01 cm01 at 351 nm for B f PQ. All
ligands are protonated in aqueous pH 6.0 solution; (I) at
this condition is singly charged ( /1) and (II) and (III)
are doubly charged ( /2). The protonation state of (IV)
is unknown. Changing the pH may alter the protonation
state and, hence, the spectroscopic properties. All other
chemicals were of analytical grades and used as obtained
from Sigma Chemical Co.

The optical factor depends on the angle a that the transition moment of the ligand makes with the polynucleotide
helix axis. The angle brackets denote an ensemble average over the angular distribution. The orientation factor,
which reflects the degree of orientation of the polynucleotide in the flow, is determined using the polynucleotide
dichroism at 260 nm as an internal reference, assuming
an effective angle of 867 between the p r p* transition
moments of the nucleotide bases and the polynucleotide
helix axis.29,36,37 S depends on the contour length and
flexibility of the polynucleotide, the flow rate, and the
temperature and viscosity of the medium. The polynucleotide samples were oriented using a flow Couette
cell with outer rotating cylinder. The LD spectra of the
oriented sample were obtained using a Jasco J-500A spectropolarimeter. An Oxley prism was used in order to convert the circularly polarized light into linear as described
elsewhere.38

Circular Dichroism
CD is defined as the differential absorption of left and
right circularly polarized light.
CD( l ) AL ( l ) 0 AR ( l )

(1)

Although BePI, (3 *-amino)proylamino-BePI, BgPI, and

B f PQ are achiral molecules, they may acquire CD upon
binding to chiral polynucleotides. The origin of this induced CD of an achiral ligand chromophore in a polynucleotide complex is generally believed to be a result
of nondegenerate coupling between transitions of the ligands and transitions of the surrounding chirally arranged
bases of the polynucleotide host.30,31 All CD spectra were
measured on a Jasco J-720 spectropolarimeter in a 1 cm
quartz cell.

Linear Dichroism
Flow LD at a given wavelength of light is defined as
the differential absorption of light polarized parallel and
perpendicular, respectively, to the flow direction.29
LD( l ) A\ ( l ) 0 A ( l )

(2)

Division of the LD spectrum by absorption spectrum results in a dimensionless quantity, the reduced linear dichroism (LD r ):
LD r ( l )

LD( l )
Aiso ( l )

(3)

where Aiso ( l ) is the isotropic absorption of the sample at

rest. LD r can be denoted as a product of S and O, which
are the orientation factor and the optical factor of the
sample, 29 respectively.
LD r S 1 O 3S

( 3 cos 2a 0 1)
2

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Absorption
Absorption spectra of the sample were taken by a Cary
2300 spectrophotometer using a 1 cm quartz cell.

RESULTS
Absorption
Absorption spectra of the BePI, BgPI and B f PQ
derivatives, free in solution and complexed with
poly[d(A-T)]2 , poly(dA)rpoly(dT), and poly(dA)r2poly(dT), are shown in Figure 2. The absorbances of the uncomplexed DNAs have been
subtracted in the DNA region to give an estimate
of the effect of DNA binding on the ligand spectra.
The absorption spectra illustrated in fact include
contributions in addition to that of the bound ligand:
the absorbance of any free ligand and also any
change in the DNA absorption due to ligand binding
contribute to the spectrum illustrated.
Upon binding to all polynucleotides, the BePI
ligands exhibit 5 nm red shifts (from 362 nm to
367368 nm) and about 3040% hypochromism
(the effect being slightly larger for I-BePI than for
the II-BePI complexes). In the BgPI case, whereas
nearly 50% hypochromism and 5 nm red shift is
observed in the DNA absorption region, a red shift
of 5 nm and no hypochromism is observed around
350 nm. On the other hand, neither any hypochromism nor a red shift is seen below 350 nm for
B f PQ. When B f PQ is bound to the poly(dA)r2poly(dT) triplex, a small hypochromism above 350
nm is apparent. Interestingly, when B f PQ is complexed with the duplexes, a hyperchromism and blue
shift is observed [Figure 2(d)].
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105

FIGURE 2 Absorption spectrum of (a) 3-methoxy-11-amino-BePI, (b) 3-methoxy-11-[(3 *amino)propylamino]-BePI, (c) 3-methoxy-7-[(3 *-diethylamino)propylamino]BgPI, and (d)
3-methoxy-11-[(3 *-amino)propylamino]B f PQ complexed with poly[d(A-T)]2 (dotted
curve), poly(dA)rpoly(dT) (dashed curve), and poly(dA)r2poly(dT) triplex (solid curve).
[BePI] 11 mM, [other ligands] 10 mM, [polynucleotide] 50 mM in adenine base.
Absorption spectrum of corresponding polynucleotide has been subtracted. The unbound ligand
absorption spectra is denoted by a thin trace in each panel.

LD and LD r
Figure 3 shows LD spectra of 3-methoxy-11-aminoBePI (I) and 3-methoxy-11-[(3 *-amino)propylamino]-BePI (II) complexed with poly[d(A-T)]2 ,
poly(dA)rpoly(dT), and poly(dA)r2poly(dT),
recorded with steady-state shear-flow alignment.
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The LD of the free polynucleotides are depicted for

comparison. The LD signals at all wavelengths for
all systems are negative. The LD signals of the pure
DNAs are negative as expected from the preferentially perpendicularly oriented nucleo-bases under
our experimental conditions, 29 but the signals cannot be compared directly because of differences in
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Kim et al.

FIGURE 3 LD spectrum of (a) 3-methoxy-11-amino-BePI, (b) 3-methoxy-11-[(3 *-amino)propylamino]-BePI, (c) 3-methoxy-7-[(3 *-diethylamino)propylamino]BgPI, and (d) 3-methoxy-11-[(3 *-amino)propylamino]B f PQ complexed with poly[d(A-T)]2 (dotted curve),
poly(dA)rpoly(dT) (dashed curve), and poly(dA)r2poly(dT) triplex (solid curve). Sample
concentrations are the same as in Figure 2. The thin traces in (a) are the LD of the polynucleotides, poly[d(A-T)]2 (dotted), poly(dA)rpoly(dT) (dashed), and poly(dA)r2poly(dT) triplex (solid).

lengths and flexibility between the polynucleotides,

providing different values of the orientation factor
S, of Eq. (4). The possibility of a minor groove
binding mode, which would have given a positive
LD signal for any ligand transitions polarized near
the long axis of the molecule, can be immediately
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excluded because of the negative ligand LD signals

for all ligand transitions.
The LD r spectra for the complexes, calculated
using Eq. (3), are shown in Figure 4. The LD r
spectra of the free polynucleotides are also depicted
in Figure 4(a) for comparison. The addition of each
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107

FIGURE 4 Calculated LD r of the (a) 3-methoxy-11-amino-BePI, (b) 3-methoxy-11-[(3 *amino)propylamino]-BePI, (c) 3-methoxy-7-[(3 *-diethylamino)propylamino]BgPI, and (d)
3-methoxy-11-[(3 *-amino)propylamino]B f PQ complexes. Concentrations are the same as in
Figure 2. A-T, A : T, and TAT denote, respectively, poly[d(A-T)]2 , poly(dA)rpoly(dT),
and poly(dA)r2poly(dT) triplex. The LD r of ligand free polynucleotides are shown in (a) as
thin curves.

ligand to the alternating duplex poly[d(A-T)]2

causes a significant increase in the magnitude of the
DNA LD r ; the LD r signals in the ligand region for
the complexes with this polynucleotide are fairly
flat and slightly larger in magnitude than in the DNA
region. In both absorption regions the LD r is also
essentially wavelength independent, consistent with
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a perpendicular orientation (907 from the helix axis)

for all transition moments. These observations are
all in accord with the ligands binding intercalatively
and lengthening and stiffening the helix. The LD r
for the poly[d(A)]rpoly[d(T)] complexes show
little change in orientation of the DNA. The negative amplitude of the LD r in the nucleo-base absorpbpal

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tion region is close to that displayed by the long

wavelength transitions of the BePI chromophore for
all of the complexes, again indicating that the transition dipoles of the polynucleotide bases and of the
BePI chromophore have the same angle with respect
to the helix axis. With the triplex, a 15% decrease
of LD r in the DNA region is observed for the BePI,
BgPI, and B f PQ complexes. In all except the 3methoxy-11-amino-BePI complex, the ligand LD r
is larger than that in the DNA region.

small positive one from about 290 to 350 nm. A

broad negative band with more structure than the
corresponding normal absorption is evident from
250 to 280 nm.
The ICD of the two duplexes with 3-methoxy11-[(3 *-amino)propylamino]B f PQ (IV) resemble one another, with the nonalternating duplex ICD
being almost the mirror image of its form with (II).

DISCUSSION
Circular Dichroism
The induced CD (ICD CD of DNA plus ligand
complex minus DNA CD) spectra of the benzopyridoindole and benzopyridoquinoxaline ligands in the
polynucleotide complexes are shown in Figure 5.
Perturbations to the DNA could be contributing to
the ICD in the DNA region of the spectrum (below
290 nm); however, in contrast to the absorption
data, the achiral free ligands do not affect the ICD.
The ICD observed for ligand-polynucleotide complexes depend in a sensitive but complicated way on
the environment and local orientation of the bound
ligand.30,31 The ICD spectrum of the complexes with
the poly(dA)rpoly(dT) duplex and the poly(dA)r2poly(dT) triplex are similar but are at somewhat
longer wavelengths for the 300 nm negative band
of (I), (II), and (III). The ICD spectra of (IV)
with the two polynucleotides are quite different.
At 370 nm there is no evidence of an ICD signal
in the 3-methoxy-11-amino-BePI (I) complexes, in
contrast to the situation for the 3-methoxy-11-[(3 *amino)propylamino]-BePI complexes. The 300 nm
negative band of the nonalternating complexes is
the first of four bands of alternating signs that are
almost excitonic in appearance. The signs in the
250 nm DNA region are the same for all complexes.
The 250 nm part of this series is likely to be dominated by exciton coupling with the DNA base transitions.
The 3-methoxy-11-[(3 *-amino)propylamino]BePI (II) complexes all have a small positive band
at 370 nm; at 330 nm the nonalternating DNAs also
induce a positive CD into the ligand transitions,
whereas that induced by the alternating DNA is
negative. The ICD of the poly[d(A-T)]2 complex
is smaller than for the analogous 3-methoxy-11amino-BePI complex, the two bands at 280 nm and
300 nm canceling rather than reinforcing.
The ICD of 3-methoxy-11-amino-BgPI (III)
with poly[d(A)]rpoly[d(T)] and with poly[d(A)]rpoly[d(T)]2 looks similar to that of the other
complexes. With poly[d(A-T)]2 , however, there is
a very small negative signal at 360 nm and a very
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Linear Dichroism
The negative LD observed for the ligand transitions
of all the complexes immediately excludes the possibility of a minor groove binding mode.29 Furthermore, the negative LD r values in the nucleobase
absorption (around 260 nm) and ligand absorption
(300400 nm) regions are of similar magnitude for
all the complexes examined, indicating that the
planes of the nucleobases and that of the ligand
molecules lie parallel to each other. By comparison
with the LD r amplitudes at 260 nm of the uncomplexed duplex polynucleotides, it is also inferred
that the base planes remain essentially perpendicular
to the helix axis of polynucleotide upon ligand binding. The decrease of the LD r amplitude at 260 nm
observed for the triplex complexes could indicate
a minor change of base tilt or a slightly reduced
persistence length of this structure upon interaction
with the ligands. The converse is true for the poly[d(A-T)]2 complexes.

Circular Dichroism
The ICD spectra of the electric dipole-allowed transitions of the benzopyridoindole molecules complexed with poly(dA)rpoly(dT) and triplex poly
(dA)r2poly(dT) are approximately mirror images
of that of the poly[d(A-T)]2 complexes outside the
DNA region of the spectrum. The duplex complexes
with 3-methoxy-11-amino-BePI have mirror image
ICD spectra across the whole wavelength range.
This leads to two questions, the first being why the
mirror image is found in the ligand region and the
second is why for two of the ligands the DNA region
is approximately independent of the identity of the
DNA yet for the other two it is not.
Although the sign of the ICD of an intercalator
depends not only on its orientation in the intercalation pocket but also on the identity of the site, 31,36
it is often the case that the ligand ICD signals induced by poly[d(A-T)]2 and poly(dA)rpoly(dT)
are very similar. Hence, one might speculate that
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FIGURE 5 ICD of the (a) 3-methoxy-11-amino-BePI, (b) 3-methoxy-11-[(3 *-amino)propylamino]-BePI, (c) 3-methoxy-7-[(3 *-diethylamino)propylamino]BgPI, and (d) 3-methoxy11-[(3 *-amino)propylamino]B f PQ complexes. Samples are the same as in Figure 2. A-T,
A : T, and TAT denote, respectively, the poly[d(A-T)]2 , poly(dA)rpoly(dT), and poly(dA)r2poly(dT) triplex.

the ligands (I), (II), and (III) are rotated 907 in

the intercalation pocket of poly[d(A-T)]2 compared to their orientation with poly(dA)rpoly(dT)
and poly(dA)r2poly(dT). Alternatively, it is possible that the CD arises from a skewed conformation
of 3-methoxy-11-amino-BePI and 3-methoxy-11[(3 *-amino)propylamino]-BePI ligands themselves when they are bound to DNA. A preference
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for different enantiomers by the alternating and nonalternating DNAs could then account for the oppositely signed CDs. If this were the case, the similarity of the nonalternating ICD spectra for the BePI
complexes with those for 3-methoxy-11-aminoBgPI would require BgPI to be similarly skewed.
However, the very small ligand-region ICD for the
complex of this compound with poly[d(A-T)]2 rebpal

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Kim et al.

quires it to have retained a planar geometry. Such

a difference between the ligands is less likely than
that all three ligands bind similarly in planar geometries. The longer structure of 3-methoxy-11-aminoBgPI, however, allows it only a 457 rotation
[rather than 907 allowed for (I) and (II)] in the
intercalation site with the alternating polymer and
hence the observed small ICD.
As for the difference in poly[d(A-T)]2 spectra
observed in the DNA region for the different ligands, it may simply be that the ligand absorption
maximum in this region is shifted to slightly longer
wavelength so that the energy difference in exciton
coupling of the ligand transitions with those of the
DNA bases is reversed.

CONCLUSIONS
We conclude that, based on LD r , the planar polycyclic parts of both 3-methoxy-11-amino-BePI and 3methoxy-11-[(3 *-amino)propylamino]-BePI
as
well as those of the BgPI and B f PI derivatives are
intercalated between the base pairs of poly[d(AT)]2 and poly(dA)rpoly(dT) as well as between
the base triplets of poly(dA)r2poly(dT). It is also
interesting to note that both the nonalternating duplex and the triplex induce similar CD spectra into
the ligand transitions of the BePI and BgPI derivatives suggesting their binding modes are very similar. By contrast, the ligand-region induced CD for
the BePI complexes with poly[d(A-T)]2 is almost
a mirror image of that found with the nonalternating
DNAs. This suggests that the orientation of these
ligands is different within the intercalation pockets
in the alternating and nonalternating DNAs. The
longer BgPI complex has a small ligand ICD with
the alternating duplex that is also opposite in sign
from the larger signals with the nonalternating
DNAs. It may therefore be concluded that the BgPI
is rotated in the alternating duplex intercalation
pocket by a smaller amount from the nonalternating
binding geometry than are the other two ligands.
The ICD of B f PI with the triplex resembles that
of the other ligands so its binding mode may be
concluded to be similar and distinct from that observed for both duplexes.

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