Supramolecular assemblies of DNA with neutral nucleoside amphiphiles

Article abstract of DOI:10.1039/b412670j

A neutral uridine-based amphiphile is described which condenses plasmid DNA. AFM studies show that the three distinct structural components of the amphiphile (i.e, nucleobase, alkyl chains, and poly(ethylene glycol)) are required for the formation of DNA-

Full text of DOI:10.1039/b412670j

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Supramolecular assemblies of DNA with neutral nucleoside
amphiphiles{
Philippe Barthelemy,a Carla A. H. Prata,b Shaun F. Filocamo,b Chad E. Immoos,d Benjamin W. Maynor,e
S. A. Nadeem Hashmi,f Stephen J. Leec and Mark W. Grinstaff*b
Received (in Columbia, MO, USA) 17th August 2004, Accepted 17th December 2004
First published as an Advance Article on the web 21st January 2005
DOI: 10.1039/b412670j
range.21,22 Micelles are formed with amphiphiles 1 and 2 with
CMC values of 6.4 6 1025 and 4 6 1024 M, respectively. The
A neutral uridine-based amphiphile is described which con-
denses plasmid DNA. AFM studies show that the three distinct
lower CMC observed with amphiphile 1 compared to 2 is
consistent with amphiphile 1 possessing increased intermolecular
interactions.
structural components of the amphiphile (i.e, nucleobase, alkyl
chains, and poly(ethylene glycol)) are required for the forma-
tion of DNA–amphiphile supramolecular assemblies on a mica
surface.
The ability of compounds 1 through 5 to condense plasmid
DNA on a mica surface was investigated using atomic force
microscopy (AFM). DNA is well known to undergo an elongated
coil-to-globular transition in the presence of amphiphilic cationic
polymers. Several AFM studies have described the toroidal
structures formed in DNA–polymer condensates.23,24 Fig. 2 shows
the atomic force micrographs of plasmid DNA (pBR322 plasmid
DNA E. coli strain RII) in the presence of 1 and 2. All the studies
were performed above a concentration of 5 6 1024 M (i.e., above
the CMCs for the amphiphiles). Only amphiphile 1 condenses
plasmid DNA. The micrographs at this amphiphile concentration
(1 mg mL21) show more than 80% of the structures on the surface
are of a collapsed state. The AFM images of DNA in the presence
of 3, 4, or 5 are very similar to the micrographs of 2 or DNA
without an amphiphile. Upon examining a series of micrographs
of DNA (0.5 mg mL21) with varying amphiphile 1 concentrations
from 0.5 to 5 mg mL21 we observed a range of collapsed states
from the partial collapse to the condensed supramolecular
structure, a toroid. The AFM image of a toroid at higher
magnification is shown in Fig. 3. The toroid has an outer and inner
diameter of approximately 100 nm (av. 106 ¡ 22 nm) and 20 nm
(av. 21 ¡ 11 nm), respectively, and is similar to structures
previously observed with cationic DNA condensation agents.24
Under similar conditions toroids are not observed with compound
Amphiphiles play important roles in diverse applications including
building blocks for nanotechnology, tools for biophysical mem-
brane studies, and carriers for drug delivery.1–13 The supramole-
cular structures formed in those systems are a consequence of
non-covalent interactions (e.g., electrostatic, hydrophobic,
H-bonding, etc.) and vary in size from a few nanometers to
millimeters. We are interested in determining the molecular design
criteria required for an amphiphile to have selective interactions
with biological macromolecules; such systems are likely to advance
our understanding, ability to manipulate biological systems, and
capability to form supramolecular assemblies. For example, the
binding of cationic amphiphiles with biological macromolecules,
such as DNA, to form assemblies is of widespread interest and
clinical importance.14–17 However, the electrostatic interaction
between the cationic amphiphile and anionic DNA is non-specific
and therefore these amphiphiles can also bind a number of
proteins at pH = 7.4, since many proteins have a low isoelectric
point (e.g., albumin). Herein, we describe a neutral nucleoside
amphiphile for condensing plasmid DNA on a surface.
The design principle of this neutral amphiphile relies on
cooperative non-covalent interactions, such as the molecular
forces that hold nucleic acid helices together (Watson–Crick/
Hoogsteen hydrogen-bonding and base p-stacking), and hydro-
phobic chain–chain interactions present in lipid bilayers.18–20
Consequently, this amphiphile possesses a DNA nucleobase, a
hydrophobic region (e.g., alkyl chains), and a poly(ethylene glycol)
(PEG) segment as shown in Fig. 1 (structure 1). Amphiphiles 1–4
were synthesized and studied to determine the possible role of and
interplay between the nucleobase, alkyl chains, and PEG of the
amphiphile structure in the formation of DNA–amphiphile
assemblies (Fig. 1).{
Given the amphiphilic structure of these compounds, it is likely
that micelles will form in aqueous solution. Previous results with
PEG derivatized phospholipids showed spherical micelle forma-
tion with critical micelle concentrations (CMCs) in the mM
{ Electronic supplementary information (ESI) available: detailed experi-
mental information for the synthesis of the amphiphiles. See http://
www.rsc.org/suppdata/cc/b4/b412670j/
Fig. 1 Nucleoside amphiphiles and structural analogs.
Chem. Commun., 2005, 1261–1263 | 1261
*mgrin@bu.edu
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DNA is not observed with 4 or 5, highlighting the significance of
the hydrophobic chains. Finally, toroid formation is not observed
with 3 or 5, indicating the need for a large molecular weight PEG
chain.
All three structural components in the amphiphile (i.e,
nucleobase, alkyl chain, and PEG) are important for DNA
condensation. For example, the binding constant for just a single
U–U base–base interaction in water is small. In a non-hydrogen
bonding organic solvent (e.g., CHCl3), the binding constant is
estimated to be less than 102 M21 and orders of magnitude weaker
than known cationic amphiphiles like DOTAP (1,2-dioleoyloxy-3-
(trimethylammonio)propane; 107 M21) binding to DNA in
water.25,26 Thus, it is unlikely that the nucleobase itself is
responsible for DNA condensation, consistent with the experi-
mental results – compound 1 but not 5 affords toroids. The
hydrophobic alkyl chain–chain interactions play a significant role,
as observed in liposomes,20,27 and contribute to favorable enthalpic
energies for the condensation, given that 1 but not 4 gives toroids.
The role of the high molecular weight PEG chain is multifaceted.
First, the PEG increases the limited solubility of the hydrophobic,
long-chain acylated nucleoside in aqueous solution. Second, the
PEG and alkyl chains are required for micelle formation. Third, it
has been shown that very high concentrations of PEG in aqueous
solution will precipitate DNA.28 This effect is manifested through
a loss of water hydration of the DNA via increased interactions
between the DNA and PEG. The PEG concentrations used in our
experiments are significantly lower, and DNA condensation is not
observed with PEG 5000 Mw under these experimental conditions.
Thus, we propose that 1 condenses plasmid DNA through an
organized macromolecular crowding mechanism.29,30 In solution,
the micelles composed of the uridine amphiphiles interact with
each other as well as with the plasmid DNA, and condensation
occurs when the local micelle concentration becomes high near the
DNA. This is consistent with the observation that amphiphiles 1
and 2 form micelles, but only 1, the amphiphile that possesses the
uridine, condenses DNA.
Fig. 2 AFM images of plasmid DNA in the presence of (top) amphiphile
1 and (bottom) amphiphile 2.
In summary, a non-cationic amphiphile for the condensation of
DNA is described. With the objective of mimicking a nucleosome,
a recent report describes a glycocluster amphiphile which binds
DNA through interactions with the phosphate backbone.7 These
results provide further evidence for the use of non-catonic
interactions to manipulate DNA. The importance of multiple
non-covalent weak interactions working in unison to form
supramolecular structures cannot be overstated. The development
of amphiphiles possessing affinity to a biological macromolecule
(e.g., protein, mRNA, DNA, etc.) through more specific interac-
tions, as opposed to more general electrostatic forces, will broaden
their use in the areas of biological chemistry, biomaterials, and
biotechnology.
This work was supported by the Army Research Office
(DAAD19-02-1-0386). CEI thanks the NRC Research
Associateships Program for a postdoctoral fellowship.
Fig. 3 3D AFM image of a toroid formed between amphiphile 1 and
DNA at higher magnification (outer diameter of the toroid is
approximately 100 nm). Image is a 300 nm2 with a z range of 10 nm.
Philippe Barthelemy,a Carla A. H. Prata,b Shaun F. Filocamo,b
Chad E. Immoos,d Benjamin W. Maynor,e S. A. Nadeem Hashmi,f
Stephen J. Leec and Mark W. Grinstaff*b
2 which lacks the nucleobase, confirming the requirement of the
uridine in the amphiphile structure. Since the uridine but not
the OMe analog condenses DNA, it is reasonable to propose
that the nucleobase weakly interacts with the plasmid DNA via
H-bonding and/or p-stacking interactions. The condensation of
aFaculte´ des sciences d’Avignon, 33 rue Louis Pasteur, F-84000,
Avignon, France
bDepartments of Chemistry and Biomedical Engineering, Boston
University, Boston, MA 02215, USA. E-mail: mgrin@bu.edu;
Fax: +1-617-353-6466; Tel: +1-617-358-3429
1262 | Chem. Commun., 2005, 1261–1263
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cArmy Research Office, RTP, NC 27709, USA
dDepartment of Chemistry, California Polytechnic State University,
San Luis Obispo, CA 93407, USA
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eDepartment of Chemistry, University of North Carolina at Chapel Hill,
NC 27599, USA
fDepartment of Chemistry, Duke University, Durham, NC 27708, USA
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