ScienceWeek

SCIENCEWEEK

ScienceWeek
September 19, 2003
Vol. 7 Number 38A

An Online Digest of Research in the Sciences

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There is no national science
just as there is no national multiplication table;
what is national is no longer science.
-- Anton Chekhov (1860-1904)

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Section 1

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Part A - Symposium: Polymer Electrolytes

1. Introduction
2. On Zwitterionic Polymers
3. Ionic Conductivity in Crystalline Polymer Electrolytes
4. Protein Folding and Hydrophobic Clusters
5. Like-Charge Attraction Between Polyelectrolytes Induced by
Counterion Charge Density Waves
6. Long-Range Interactions Within a Nonnative Protein
7. Biosensors from Conjugated Polyelectrolyte Complexes
8. Molecular Basis for Exponential Growth of Polyelectrolyte
Multilayers
9. Probing Protein Electrostatics with a Synthetic Fluorescent
Amino Acid
10. On Charge Inversion in Polymers

Notices and Subscription Information

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Section 2

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1. INTRODUCTION

Of all materials used by man, those based on polymer molecules
have been around longest. Wood, for example, was pressed into
service long before metals were being smelted, and wood is
composed of cellulose, lignin, and resins, all of which are
natural organic polymers. Nevertheless, an adequate knowledge of
polymer structure and properties has been acquired only during
recent decades. The reason for this delay lies in the fact that
these molecules are frequently very large and complex compared
with anything encountered in metals and ceramics. Analysis of
such huge aggregates of atoms, with molecular weights frequently
running into the hundreds of thousands, has required the
sophistication of modern instrumentation, and polymer science is
still a relatively young discipline.

The etymology of the word "polymer" is straightforward enough.
The Greek word "poly" means many, and another Greek word "meros"
means parts. The choice of this name stems from what has been
found to be the defining characteristic of polymers; they are
formed by the coming together of many smaller units. Some of
these extend thread-like in one dimension, while others form
sheets, and still others are arranged in three-dimensional
networks. The individual units are called "monomers", and most
common polymers are composed of regular repetitions of just a few
monomers, and often only one.

There is no formal restriction on the composition of a polymer,
and examples like asbestos are not even organic. Synthetic
polymers of the organic type are usually associated with the word
"plastic", a term which is somewhat misleading in that it refers
to a material which is not plastic in the physical sense. A
plastic is best defined, therefore, as a material based primarily
on polymeric substances, which is stable in normal use, but which
at some stage during its manufacture is physically plastic.

The early efforts in the field of organic polymers were of
necessity directed towards naturally occurring polymers, but once
the regularity of their structures had been revealed, chemists
began to tailor-make artificial polymers by stringing together
combinations of monomers that do not occur naturally. This
activity has now mushroomed into the vast industry that annually
produces millions of metric tons of plastics, synthetic fibers,
and related products.

In contrast to the technologies of metals and ceramics, the
origins of which are antique and obscure, the mileposts of
development in the polymer industry are so recent that the story
is well documented. The first polymeric material to attract
scientific interest appears to have been silk. Robert Hooke
(1635-1703) noted, in 1665, that this substance appears to be a
type of glue, and he suggested that the product of the silkworm
could be imitated by drawing a suitable glue-like material out
into a thread. It transpired that he was more than two hundred
years before his time.

Not surprisingly, the earliest technological use of materials of
this type, in a rudimentary polymer chemistry, took advantage of
naturally occurring substances. Among these were amber, rosin,
gum arable, asphalt, and pitch, the latter two being valued for
their waterproofing properties as early as five thousand years
ago. Natural resins were familiar to the early Egyptians who used
them in varnishing sarcophagi. Shellac, which was in common use
by the middle of the third century AD, is unique among the resins
in that it is of animal origin. It is secreted by the insect
Tachardia lacca which is found on the lac tree Butea frondosa.
Shellac is still used, alcoholic solutions having a consistency
convenient for varnishes, polishes, electrical insulation, and,
at greater concentration of the polymer, sealing wax.

Early explorers of the Central Americas were amazed by a
substance used by the native Indians both for ball games and in
the production of water-resistant boots. This elastic gum, now
known as rubber and obtained primarily from the tree Hevea
brasiliensis, was first made the object of scientific study in
1751, by Charles-Marie de La Condamine. Joseph Priestley (1733-
1804) must be given the credit for discovering its utility as an
eraser of pencil marks, while the raincoat made of rubberized
fabric is synonymous with the surname of its developer, Charles
Mackintosh.

Adapted from: Rodney Cotterill: The Cambridge Guide to the
Material World. Cambridge University Press 1985, p.215. More
information at:
http://www.amazon.com/exec/obidos/ASIN/0521246407/scienceweek

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2. ON ZWITTERIONIC POLYMERS

An "amphoteric" molecule is a molecule that can react as acidic
towards strong bases and as basic towards strong acids (e.g., an
amino acid). An "ampholyte" is any amphoteric electrolyte, and a
"polyampholyte" is a polymer with ampholyte properties. Proteins
are polyampholytes.

The following points are made by A.B. Lowe and C.L. McCormick
(Chem. Rev. 2002 102:4177):

1) Polymers containing ionic groups are among the most important
classes of macromolecules. These types of polymers range from
naturally occurring biopolymers such as proteins and nucleotides
to synthetic viscosifiers and soaps. Ionic polymers may be
divided into two groups, polyelectrolytes(1-4) and
polyzwitterions. Polyelectrolytes contain anionic or cationic
groups, while polyzwitterions contain both anionic and cationic
groups. Polyzwitterions have a wide variety of applications that
include ion exchange, chelation to bind trace metals (Hg, Cd, Cu,
and Ni) from drinking water, sewage treatment, soil conditioning,
paper reinforcement, pigment retention, and formulation in
shampoos and hair conditioners.(5)

2) A characteristic of polyelectrolytes is chain extension, and
thus large hydrodynamic volume, in deionized water at low
concentrations. This is due to Coulombic repulsions between
charged groups along the polymer chain, forcing the polymer into
an extended rodlike conformation. The addition of low molecular
weight electrolyte or changes in solution pH screens the
repulsive electrostatic forces, and the polymer coil shrinks,
adopting a more entropically favored conformation. This is known
as the "polyelectrolyte effect".

3) For polyzwitterions, the charges may be located either on the
pendent side chains of different monomer units or the same
monomer unit, or in the case of some polyesters,
polyphosphazenes, and polyphosphobetaines, one or both of the
charges may be located along the polymer backbone.

4) The solution behavior of polyzwitterions is often opposite
that of polyelectrolytes, exhibiting the so-called
"antipolyelectrolyte effect". Chain expansion occurs upon the
addition of low molecular weight electrolyte, although this is
very much dependent upon chemical structure, composition, and
solution conditions.

5) Polyampholytes are interesting for numerous reasons, not the
least of which is the fact that they are synthetic analogues of
naturally occurring biological molecules such as proteins and
find applications in areas such as lithographic film, emulsion
formulations, and drag reduction. The structure-property
relationships of polyampholytes are dictated by Coulombic
attractions between anionic and cationic species on different
monomer units. The response in aqueous solution exhibited by
polyampholytes is very much dependent upon both the chemical
structure and composition of the polymer. There exist four
subclasses of polyampholytes based on their responses to changes
in pH as described previously. First, the polyampholyte may
contain both anionic and cationic species that may be
neutralized. Second, the anionic group may be neutralized while
the cationic species is insensitive to changes in pH. Third, the
cationic species may be neutralized while the anionic species is
insensitive to changes in pH. Finally, both the anionic and
cationic species may be insensitive to changes in solution pH
over the useful range.

References (abridged):

1. Dubin, P.; Bock, J.; Davies, R. M.; Schuiz, D. N.; Thies, C.
Macromolecular Complexes in Chemistry and Biology; Spnnger-
Verlag: Berlin, 1994.

2. Mortimer, D. A. Polym. Int. 1991, 25, 29.

3. Bromberg, L. In Handbook of Surfaces and Interfaces of Materi-
als, Nalwa, H. S., Ed.; Academic Press: New York, 2001; Vol 4.,
Chapter 7.

4. Oosawa, F. Polyelectrolytes; Marcel-Dekker: New York, 1971.

5. Salamone, J. C.; Rice, W. C. Encyclopedia of Polymer Science
and Engineering, 2nd ed.; Wiley-Interscience: New York, 1988;
Vol. 11, p 514.

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3. IONIC CONDUCTIVITY IN CRYSTALLINE POLYMER ELECTROLYTES

The following points are made by Z. Gadjourova et al (Nature 2001
412:520):

1) Polymer electrolytes are the subject of intensive study, in
part because of their potential use as the electrolyte in all-
solid-state rechargeable lithium batteries(1). These materials
are formed by dissolving a salt (for example LiI) in a solid host
polymer such as poly(ethylene oxide) (2-5), and may be prepared
as both crystalline and amorphous phases. Conductivity in polymer
electrolytes has long been viewed as confined to the amorphous
phase above the glass transition temperature, T(subg), where
polymer chain motion creates a dynamic, disordered environment
that plays a critical role in facilitating ion transport(2,3).

2) Polymer electrolytes form crystalline compounds at certain
discrete compositions. For example, the crystalline compound
PEO3:LiCF3SO3 is obtained by dissolving LiCF3SO3 in poly(ethylene
oxide) (PEO) with a composition corresponding to three ether
oxygens per lithium ion. Polymer electrolytes may also be
obtained as amorphous materials over a wide range of compositions
and temperatures, including, in some cases, the compositions at
which crystalline complexes exist. Ionic conductivity occurs in
the amorphous phase above T(subg), where ion transport is induced
by local motion of polymer chain segments repeatedly creating new
coordination sites into which the ions may then migrate. For
example, NMR studies of ionic motion within the PEO:LiCF3SO3
system, at compositions where the crystalline 3:1 complex and the
amorphous phase coexist above T(subg), demonstrate that ion
transport occurs in the amorphous phase.

3) The authors demonstrate that in contrast to the prevailing
view, ionic conductivity in the static, ordered environment of
the polymer crystalline phase can be greater than that in the
equivalent amorphous material above T(subg). Moreover, the
authors demonstrate that ion transport in crystalline polymer
electrolytes can be dominated by the cations, whereas both ions
are generally mobile in the amorphous phase. Restriction of
mobility to the lithium cation is advantageous for battery
applications. The authors suggest the realization that order can
promote ion transport in polymers is interesting in the context
of electronically conducting polymers, where crystallinity
favours electron transport.

References (abridged):

1. Croce, F., Appetecchi, G. B., Persi, L. & Scrosati, B.
Nanocomposite polymer electrolytes for lithium batteries. Nature
394, 456-458 (1998)

2. Gray, F. M. Polymer Electrolytes (RSC Materials Monographs,
The Royal Society of Chemistry, Cambridge, 1997)

3. Scrosati, B. (ed.) Applications of Electroactive Polymers
(Chapman & Hall, London, 1993)

4. Bruce, P. G. (ed.) Solid State Electrochemistry (Cambridge
University Press, Cambridge, 1995)

5. Fenton, D. E., Parker, J. M. & Wright, P. V. Complexes of
alkali metal ions with poly(ethylene oxide). Polymer 14, 589
(1973)

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4. PROTEIN FOLDING AND HYDROPHOBIC CLUSTERS

The following points are made by Robert L. Baldwin (Science 2002
295:1657):

1) It has been hotly debated whether a hydrophobic collapse
precedes or occurs concomitantly with formation of secondary
structures at the beginning of protein folding (1). The role of
secondary structures in guiding the folding pathway is readily
understood because they provide the framework for the final
native structure. It has also long been recognized (2) that
burial of nonpolar (hydrophobic) side chains out of contact with
water provides the major source of the free energy change that
drives folding. But compelling models have been lacking for how
this burial might produce a "hydrophobic collapse" that initiates
folding.

2) The most plausible model has been the "hydrophobic zipper"
(3), in which clusters of nonpolar side chains stabilize
secondary structures such as a helices or beta hairpins. Examples
are known in which such hydrophobic clusters guide the folding
process (4,5). The clusters persist until folding is complete and
can be visualized in the native structure of proteins.
Hydrophobic patches have also been observed in native protein
structures at sites where two a helices interact, and there is
good evidence that hydrophobic clusters of this kind can guide
the folding process.

3) In these examples, hydrophobic clusters and secondary
structures are formed concomitantly and produce native-like
structures. But hydrophobic clusters have also been found in
denatured proteins, under conditions where secondary structures
are unstable. Some of these clusters must reflect nonnative
interactions because they cannot be found in the structures of
the native proteins. It remains unclear how these hydrophobic
clusters affect the folding process.

4) Hydrophobic clusters in denatured proteins can be detected by
various nuclear magnetic resonance (NMR) probes, including the
nuclear Overhauser effect and chemical shifts of side chain
protons. Clusters are formed not only in water but also in the
presence of the denaturant urea. A particularly useful probe for
examining which residues form a hydrophobic cluster in a
denatured protein is the transverse relaxation rate, R-2, of a
residue's amide group in the peptide backbone, measured by 15N-1H
heteronuclear NMR. The value of R-2 expected for a structureless
"random coil" can be read from the nearly horizontal plot of R-2
versus residue number. Residues forming a hydrophobic cluster
have elevated values that are readily observed in this plot.

References (abridged):

1. S. Akiyama et al., Proc. Natl. Acad. Sci. U.S.A. 99, 1329
(2002)

2. W. Kauzmann, Adv. Protein Chem. 14, 1 (1959)

3. K. A. Dill, K. M. Fiebig, H. S. Chan, Proc. Natl. Acad. Sci.
U.S.A. 90, 1942 (1993)

4. K. Zdanowski, M. Dadlez, J. Mol. Biol. 287, 433 (1999)

5. M. E. Hodsdon, C. Frieden, Biochemistry 40, 732 (2001)

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5. LIKE-CHARGE ATTRACTION BETWEEN POLYELECTROLYTES INDUCED BY
COUNTERION CHARGE DENSITY WAVES

The following points are made by T.E. Angelini et al (Proc. Nat.
Acad. Sci. 2003 100:8634):

1) Electrostatics in aqueous media is commonly understood in
terms of screened Coulomb interactions, where like-charged
objects, such as polyelectrolytes, always repel. These intuitive
expectations are based on mean field theories, such as the
Poisson–Boltzmann formalism, which are routinely used in colloid
science and computational biology.

2) Like-charge attractions, however, have been observed in a
variety of systems. Intense theoretical scrutiny over the last 30
years suggests that counterions play a central role, but no
consensus exists for the precise mechanism.

3) The authors report they have directly observed the
organization of multivalent ions on cytoskeletal filamentous
actin (a well defined biological polyelectrolyte) by using
synchrotron x-ray diffraction and discovered an unanticipated
symmetry-breaking collective counterion mechanism for generating
attractions. Surprisingly, the counterions do not form a lattice
that simply follows actin's helical symmetry; rather, the
counterions organize into "frozen" ripples parallel to the actin
filaments and form one-dimensional (1D) charge density waves.
Moreover, this 1D counterion charge density wave couples to twist
distortions of the oppositely charged actin filaments.

4) This general cooperative molecular mechanism is analogous to
the formation of polarons in ionic solids and mediates
attractions by facilitating a "zipper-like" charge alignment
between the counterions and the polyelectrolyte charge
distribution. The authors suggest these results can fundamentally
impinge on our general understanding of electrostatics in aqueous
media and are relevant to a wide range of colloidal and
biomedical processes.(1-5)

References (abridged):

1. Kirkwood, J. G. & Shumaker, J. B. (1952) Proc. Natl. Acad.
Sci. USA 38, 863–871

2. Oosawa, F. (1971) Polyelectrolytes (Dekker, New York). Ha, B.
Y. & Liu, A. J. (1997) Phys. Rev. Lett. 78, 1289–1292

3. Podgornik, R. & Parsegian, V. A. (1998) Phys. Rev. Lett. 80,
1560–1563

4. Lyubartsev, A. P., Tang, J. X., Janmey, P. A. & Nordenskiold,
L. (1998) Phys. Rev. Lett. 81, 5465–5468

5. Stevens, M. J. (1999) Phys. Rev. Lett. 82, 101–104

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6. LONG-RANGE INTERACTIONS WITHIN A NONNATIVE PROTEIN

The following points are made by J. Klein-Seetharaman et al
(Science 2002 295:1719):

1) Incompletely folded states of proteins are coupled to cellular
processes such as protein synthesis, translocation across
membranes, and signal transduction (1,2). In addition,
intrinsically unstructured proteins have been predicted to be
common within the genomes of all organisms (3). Unstructured and
partially folded conformations of proteins are, however, prone to
aggregate and have been implicated in a wide range of diseases
(4). The structural and dynamic characterization of nonnative
states of proteins is therefore crucial for understanding these
processes in addition to being fundamental to an understanding of
protein folding itself.

2) Nonnative states of proteins are ensembles of conformers, the
individual members of which may differ substantially in their
structural and dynamic properties. Conformational sampling of
denatured proteins can be significantly restricted, and the
existence of "compact states" has been postulated to occur (5).
In some cases, specific experimental structural information has
been obtained, although in general this information is either
indirect or highly localized. An important question relating to
all nonnative states is the extent to which long-range
interactions are important in the stabilization of nonrandom
interactions.

3) A wide range of approaches has been developed to characterize
nonnative states of proteins in atomic detail by NMR
spectroscopy, and evidence for the presence of residual structure
even under strongly denaturing conditions has been presented.
Residual structure appears to reside predominantly in hydrophobic
clusters, in which tryptophan or histidine residues are
surrounded by other hydrophobic side chains. It has been
postulated that hydrophobic clusters are stabilized by long-range
interactions and may influence the folding of the protein, for
example by acting as nucleation sites around which structure can
be formed. Hydrophobic clusters have also been identified in
nonnative states of hen lysozyme, in both the oxidized and the
reduced form in 8 M urea at pH 2 (in the reduced protein the free
sulfhydryl groups are blocked by methylation).

4) In summary: Protein folding and unfolding are coupled to a
range of biological phenomena, from the regulation of cellular
activity to the onset of neurodegenerative diseases. Defining the
nature of the conformations sampled in nonnative proteins is
crucial for understanding the origins of such phenomena. The
authors have used a combination of nuclear magnetic resonance
(NMR) spectroscopy and site-directed mutagenesis to study
unfolded states of the protein lysozyme. Extensive clusters of
hydrophobic structure exist within the wild-type protein even
under strongly denaturing conditions. These clusters involve
distinct regions of the sequence but are all disrupted by a
single point mutation that replaced residue Trp62 with Gly
located at the interface of the two major structural domains in
the native state. Thus, native-like structure in the denatured
protein is stabilized by the involvement of Trp62 in nonnative
and long-range interactions.

References (abridged):

1.  1.  P. E. Wright and H. J. Dyson, J. Mol. Biol. 293, 321
(1999)

2. S. E. Radford and C. M. Dobson, Cell 97, 291 (1999)

3. P. Romero et al., Pac. Symp. Biocomput. 3 (1998)

4. C. M. Dobson, Philos. Trans. R. Soc. London B. Biol. Sci. 356,
133 (2001)

5. B. A. Shoemaker and P. G. Wolynes, J. Mol. Biol. 287, 657
(1999)

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7. BIOSENSORS FROM CONJUGATED POLYELECTROLYTE COMPLEXES

The following points are made by Deli Wang et al (Proc. Nat.
Acad. Sci. 2002 99:49):

1) Conjugated polymers are a versatile class of organic materials
that promise utility in a variety of applications ranging from
antistatic coatings, electrodes, and transistors, to light-
emitting diodes, large area displays, photodetectors,
photovoltaic cells, and lasers (1-3). The electrical, optical,
and electrochemical properties of conjugated polymers can be
modified by chemical synthesis and are strongly affected by
relatively small perturbations, including changes in temperature,
solvent, or chemical environment. As a result of this
sensitivity, conjugated polymers are promising as sensory
materials (4,5); sensing may be accomplished by transducing
and/or amplifying physical or chemical changes into electrical,
optical, or electrochemical signals. Conjugated polymers have
been used to detect chemical species (chemosensors), such as
ions, gases (for example, trinitrotoluene), and other chemicals,
or biomolecules such as proteins, antibodies, and DNA, using
electrical, chromic, electrochemical, photoluminescent,
chemoluminescent, or gravimetric responses.

2) Contemporary biosensor and bioassay developments have focused
on mimicking natural host-receptor ("lock-and-key") interactions.
"Lock-and-key" molecular recognition can be between enzyme and
substrate, ligand and receptor, antibody and antigen, or between
two strands of nucleic acids with complementary sequences.
Although antibody-based ELISAs are widely used for detection of
biological species with high sensitivity, these assays are
relatively labor-intensive and time-consuming (hours to days) and
require two different antibodies of defined specificities to
adequately detect the target molecule (potentially making them
cumbersome to perform). Additionally, the detection of small
molecules using ELISA can seldom be accomplished because of the
recognition of one epitope by both capture and detection
antibodies. Therefore, competition assays have to be performed
that are both less accurate and more time consuming.

3) Biosensors based on conjugated polymers as sensory materials
exhibit real-time response, electrochemical or optical, to the
ligand-receptor recognition event. The coupling of a recognition
event to photoinduced electron transfer or a change in the
electronic structure of the conjugated polymer produces changes
in the luminescence, UV-visible absorption, or redox potential of
the polymer (4,5).

4) In summary: The authors report a charge neutral complex (CNC)
was formed in aqueous solution by combining an orange light
emitting anionic conjugated polyelectrolyte and a saturated
cationic polyelectrolyte at a 1:1 ratio (per repeat unit).
Photoluminescence (PL) from the CNC can be quenched by both the
negatively charged dinitrophenol (DNP) derivative, (DNP-BS), and
positively charged methyl viologen (MV2+). Use of the CNC
minimizes nonspecific interactions (which modify the PL) between
conjugated polyelectrolytes and biopolymers. Quenching of the PL
from the CNC by the DNP derivative and specific unquenching on
addition of anti-DNP antibody (anti-DNP IgG) were observed. Thus,
biosensing of the anti-DNP IgG was demonstrated.

References (abridged):

1. Skotheim, T. A., Reynolds, J. & Elsenbaumer, R. (1998)
Handbook of Conducting Polymers (Dekker, New York)

2. Friend, R. H , Gymer, R. W , Holmes, A. B , Burroughes, J. H ,
Marks, R. N , Taliani, C , Bradley, D. D. C , Dos Santos, D. A ,
Bredas, J. L , Logdlund, M., et al. (1999) Nature (London) 397,
121-128

3. McGehee, M. D., Miller, E. K., Moses, D. & Heeger, A. J.
(2000) in Twenty Years of Conducting Polymers: From Fundamental
Science to Applications, ed. Bernier, P. (Elsevier, Amsterdam)

4. Leclerc, M. (1999) Adv. Mater. 11, 1491-1498

5. McQuade, D. T., Pullen, A. E. & Swager, T. M. (2000) Chem.
Rev. 100, 2537-2574

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8. MOLECULAR BASIS EXPONENTIAL GROWTH OF POLYELECTROLYTE
MULTILAYERS

The following points are made by C. Picart et al (Proc. Nat.
Acad. Sci. 2002 99:12531):

1) The alternating dipping of a charged surface into a polyanion
and then into a polycation solution usually leads to the
progressive formation of films defined as "polyelectrolyte
multilayers" (1,2). This electrostatic self-assembly method has
been developed recently as a method for producing organic and
hybrid organic-inorganic supramolecular assemblies without
requiring extensive equipment. In particular, in the biomedical
field, these multilayers constitute versatile tools for the
design of thin films containing macromolecules such as proteins
(3), nucleic acids (4), or polypeptides with targeted properties
(5).

2) However, finding a common rule that can explain the growth
mechanism for all of the systems investigated has not been
straightforward. It is already known that polyelectrolyte
adsorption is governed by the charge reversal that appears on the
film surface after each dipping step and that constitutes the
buildup motor for the assembly. Whereas some polyelectrolyte
systems appear to grow linearly with the number of deposited
layers, others such as poly(L-lysine) (PLL)/alginate,
PLL/hyaluronan (HA), or PLL/poly(L-glutamic acid) reveal an
"exponential growth regime". This regime is characterized by film
thickness and the amount of adsorbed polyelectrolytes that
increase more rapidly than linearly with the number of deposited
layer pairs. Increasing the salt concentration in which the
buildup takes place can lead to a transition from a linear growth
regime to an exponential growth regime for the
poly(styrenesulfonate) (PSS)/poly(allylamine hydrochloride)
system or for the polydiallyldimethytlammonium chloride/PSS film
architecture.

3) In summary: The structure of poly(L-lysine) (PLL)/hyaluronan
(HA) polyelectrolyte multilayers formed by electrostatic self-
assembly was studied by the authors by using confocal laser
scanning microscopy, quartz crystal microbalance, and optical
waveguide lightmode spectroscopy. These films exhibit an
exponential growth regime where the thickness increases
exponentially with the number of deposited layers, leading to
micrometer thick films. Previously, such a growth regime was
suggested to result from an "in" and "out" diffusion of the PLL
chains through the film during buildup, but direct evidence has
been lacking. The use of dye-conjugated polyelectrolytes now
allows a direct three-dimensional visualization of the film
construction by introducing fluorescent polyelectrolytes at
different steps during the film buildup. The authors find that,
as postulated, PLL diffuses throughout the film down into the
substrate after each new PLL injection and out of the film after
each PLL rinsing, and further after each HA injection. As PLL
reaches the outer layer of the film, it interacts with the
incoming HA, forming the new HA/PLL layer. The thickness of this
new layer is thus proportional to the amount of PLL that diffuses
out of the film during the buildup step, which explains the
exponential growth regime. HA layers are also visualized but no
diffusion is observed, leading to a stratified film structure.
The authors believe that such a diffusion-based buildup mechanism
explains most of the exponential-like growth processes of
polyelectrolyte multilayers reported in the literature.

References (abridged):

1. Decher, G. (1997) Science 277, 1232-1237

2. Decher, G., Hong, J. D. & Schmitt, J. (1992) Thin Solid Films
210, 831-835

3. Ladam, G., Schaaf, P., Cuisinier, F. G. J., Decher, G. &
Voegel, J.-C. (2001) Langmuir 17, 878-882

4. Sukhorukov, G. B., Möhwald, H., Decher, G. & Lvov, Y. M.
(1996) Thin Solid Films 284-285, 220-223

5. Chluba, J., Voegel, J. C., Decher, G., Erbacher, P., Schaaf,
P. & Ogier, J. (2001) Biomacromolecules 2, 800-805

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9. PROBING PROTEIN ELECTROSTATICS WITH A SYNTHETIC FLUORESCENT
AMINO ACID

The following points are made by B.E. Cohen et al (Science 2002
296:1700):

1) High-resolution structures provide the primary basis for
analysis of protein function, yet investigations into other
physical properties such as protein dynamics and electrostatics
have been hampered by experimental limitations. For
electrostatics in particular, the lack of general techniques for
measuring static and dynamic electric fields within a protein has
made it difficult to address even a most basic question: how
polar is a protein? A protein's polarity -- that is, its capacity
to solvate charge -- affects the strengths of all electrostatic
interactions within the protein, as well as the strengths of
interactions with proteins, substrates, and other ligands, making
it a critical determinant of protein structure, stability, and,
ultimately, activity (1,2).

2) Ideas about the polarity of the protein interior have
progressed from simple models of proteins as nonpolar and
homogeneous, described by a low dielectric constant, to more
complex models that consider sources of local heterogeneity.
Computational models suggest that local dielectric properties can
vary considerably and, in some cases, correspond to polar
solvents, even in the protein interior (1,3).

3) Examining protein solvation experimentally has usually relied
on the fortuitous affinity of probes for ligand-binding sites
(4,5) or the presence of endogenous long-wavelength chromophores,
typically limiting such measurements to individual, sometimes
ill-defined, locations within particular proteins. Tryptophan
fluorescence has also been used as an intrinsic local
environmental probe, but tryptophan's utility is limited by its
complex photophysics. An ideal probe for studying protein
dynamics and electrostatics would be sensitive to its environment
and could be incorporated site-specifically throughout any
protein of interest. Toward that end, the authors have
synthesized an environment-sensitive fluorescent amino acid,
Aladan, and incorporated it site-specifically into proteins by
both nonsense suppression and solid-phase synthesis, and the
authors have used it to probe the electrostatic character of the
B1 domain of streptococcal protein G (GB1) at multiple sites by
steady-state and time-resolved fluorescence.

4) In summary: Electrostatics affect virtually all aspects of
protein structure and activity and are particularly important in
proteins whose primary function is to stabilize charge. The
authors introduce a fluorescent amino acid, Aladan, which can
probe the electrostatic character of a protein at multiple sites.
Aladan is exceptionally sensitive to the polarity of its
surroundings and can be incorporated site-selectively at buried
and exposed sites, in both soluble and membrane proteins. Steady-
state and time-resolved fluorescence measurements of Aladan
residues at different buried and exposed sites in the B1 domain
of protein G suggest that its interior of this domain is polar
and heterogeneous.

References (abridged):

1. A. Warshel and S. T. Russell, Q. Rev. Biophys. 17, 283 (1984)

2. C. N. Schutz and A. Warshel, Proteins 44, 400 (2001)

3. J. W. Pitera, M. Falta, W. F. van Gunsteren, Biophys. J. 80,
2546 (2001)

4. R. B. Macgregor and G. Weber, Nature 319, 70 (1986)

5. D. W. Pierce and S. G. Boxer, J. Phys. Chem. 96, 5560 (1992)

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10. ON CHARGE INVERSION IN POLYMERS

The following points are made by T.T. Nguyen and B.I. Shklovskii
(Phys. Rev. Lett. 2002 89:018101):

1) The inversion of the negative charge of a DNA double helix by
its complexation with a positive polyelectrolyte (PE) is used for
gene delivery. The positive charge of a DNA-PE complex
facilitates DNA contact with a typically negative cell membrane,
making penetration into the cell hundreds of times more likely
[1]. Charge inversion of DNA-PE complexes was confirmed recently
by electrophoresis [2]. At a given concentration of long DNA
helices, when the concentration of shorter PE molecules
increases, at some critical concentration the electrophoretic
mobility of a DNA-PE complex changes sign from negative to
positive.

2) The challenging and counterintuitive phenomenon of charge
inversion of a macroion by an oppositely charged PE and other
multivalent ions has attracted significant attention [3-5].
Intuitively, one can think that, when a PE completely neutralizes
a large macroion such as the DNA double helix, new molecules of a
PE do not attach to the macroion. Indeed, the Poisson-Boltzmann
approximation for the description of screening of a macroion by
any counterions including PE does not lead to charge inversion.
Charge inversion can be explained if one takes into account that
the surface potential of an already-neutralized macroion is
locally affected by a new approaching PE molecule or, in other
words, it can be explained if one takes into account correlations
between PE molecules.

3) For quantitative consideration, the charges of a macroion are
always assumed to be uniformly smeared. This approach ignores the
interference between the structure of the macroion surface and
that of a PE and clearly is not fully satisfactory. More
importantly, it is not clear whether or not charge inversion is
an artifact of the assumption of uniformly smeared charge.

4) In summary: The authors model one strand of DNA by a one-
dimensional lattice (ODL) of negative charges and consider the
problem of inversion of its charge by a positive polyelectrolyte
(PE). In the neutral state of the ODL-PE complex, each of the ODL
charges is locally compensated by a PE charge. When an additional
PE molecule is adsorbed by ODL, its charge gets fractionalized
into monomer charges of defects (tails and arches) on the
background of the perfectly neutralized ODL. Defects spread all
over the ODL, eliminating the self-energy of PE. For DNA this
fractionalization mechanism leads to a substantial inversion of
charge, a phenomenon which is widely used for gene delivery.

References (abridged):

1. A.V. Kabanov and V.A. Kabanov, Bioconjug. Chem. 6, 7 (1995);
Adv. Drug Delivery Rev. 30, 49 (1998).

2. V.A. Kabanov, A. A. Yaroslavov, and S.A. Sukhisvili, J.
Controlled Release 39, 173 (1996).

3. T. Wallin and P. Linse, J. Phys. Chem. 100, 17 873 (1996).

4. E.M. Mateescu, C. Jeppersen, and P. Pincus, Europhys. Lett.
46, 454 (1999).

5. S. Y. Park, R. F. Bruinsma, and W. M. Gelbart, Europhys. Lett.
46, 493 (1999).

--------------------------------

A NEW COLLOID STABILIZATION MECHANISM

The following points are made by V. Tohver et al (Proc. Nat.
Acad. Sci. 2001 98:8950):

1) Colloidal suspensions have widespread use in applications
ranging from advanced materials to drug delivery. By tailoring
interactions between colloidal particles, one can design stable
fluids, gels, or colloidal crystals needed for ceramics
processing, coatings, direct-write photonic devices, and
pharmaceutical applications. Long-range attractive van der Waals
forces are ubiquitous and must be balanced by Coulombic, steric,
or other repulsive interactions to engineer the desired degree of
colloidal stability.

2) The authors report the discovery of a new mechanism for
regulating the stability of colloidal particles. Colloidal
microspheres with negligible charge, which flocculate when
suspended alone in aqueous solution, undergo a remarkable
stabilizing transition upon the addition of a critical volume
fraction of highly charged nanoparticle species. Zeta potential
analysis reveals that these microspheres exhibit an effective
charge build-up in the presence of such nanoparticle species.
Reflectometry measurements, however, indicate that these
nanoparticle species do not adsorb on the microspheres under the
experimental conditions.

3) The authors therefore propose that highly charged
nanoparticles segregate to regions of near negligibly charged
microspheres because of their repulsive Coulombic interactions in
solution, and that this type of nanoparticle haloing provides a
previously unreported method for tailoring the behavior of
complex fluids.

--------------------------------

Notes:

colloid: In general, a colloid is a system of particles 1 to 1000
nanometers in diameter dispersed in another phase.

van der Waals forces: (also spelled Van der Waals) Considering
molecules that have permanent dipoles, and molecules that can
have dipoles induced by the electric fields of other molecules,
there are three possible mechanisms recognized in the formation
of the van der Waals bonds: 1) the orientation effect, in which
molecules rearrange themselves in their mutual electrical fields,
the rearrangements involving reorientations of whole molecules;
2) the static induction effect, in which molecules that are
static monopoles (ions) or dipoles may induce a static
rearrangement of the electron distribution of other molecules; 3)
the dynamic induction effect, or "dispersion" effect, in which
any molecule, polar or nonpolar, may induce in other molecules
transient electron distribution rearrangements that are time-
variant. All these mechanism involve interaction energies, and
they are "bonds" in the sense that they all involve energetic
couplings between molecules.

Coulombic force: In general, the electrical force between two
charged particles.

flocculate: In general, flocculation is the process in which
particles in a colloid aggregate into larger clumps.

Zeta potential: (electrokinetic potential) The electric potential
associated with an electrical double layer around a colloid.

Reflectometry: In general, measurement of the optical reflectance
of a surface.

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