PHYSICAL FORCES ORGANIZING BIOMOLECULES
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V.
Adrian Parsegian, PhD, Head,
Section on Molecular Biophysics Daniel
Harries, PhD, Visiting Fellow Horia
Petrache, PhD, Postdoctoral Fellow Rudi
Podgornik, PhD, Visiting Scientist |
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Despite
remarkable advances in molecular biology, the tools available to describe how
biomolecules perform their work are still relatively primitive. We attempt to
measure, characterize, and codify the forces that govern the organization of
a variety of biological molecules. Our undertaking is strengthened by its
strong connection with physical theory. Our long-term goal is to build a
practical physics of biological material through a series of measurements and
analyses of the forces governing biological molecular organization as
revealed in vivo, in vitro, and in computation. In particular,
we are working with DNA/lipid assemblies for gene therapy; DNA assemblies
such as are seen in viral capsids and in vitro; polypeptides and
polysaccharides in suspension; and lipid/water liquid-crystals. In these
systems, we simultaneously observe the structure of packing and measure
intermolecular forces or interaction energies. Ions,
lipids, and membrane-protein interactions Petrache, Podgornik;
in collaboration with Dubois, Gawrisch, Nagle, Zemb Because
ions vary widely in their effects on biological materials, ion
“specificity” beyond simple charge properties is a major issue in
biology. One overlooked property of ions is their polarizability, the ability
of the charge to shift or fluctuate, a property seen in charge fluctuation
forces. Another surprising feature of ions is their tendency to stick to
charged bilayers to an extent beyond what is expected from charge-charge
attraction. Such stickiness changes the way membranes interact and introduces
strains that can alter the way proteins are accommodated and are able to
change conformation, as in the opening and closing of transmembrane ionic
channels. Our comparisons between bilayers of phosphatidylserine (PS) and
phosphatidylcholine (PC) lipids with the same chains and at the same
temperature enable us to focus on the effects of the lipids’ headgroups
on bilayer properties. Using
X-ray diffraction and NMR spectroscopy, we found that, in the presence of
sodium ions, negatively charged PS bilayers exhibit lateral areas much
smaller than the areas of corresponding neutral PC analogs. The shrinkage
occurs despite the extra electrostatic repulsion expected for charged PS
headgroups. Such a condensation of area suggests an extra attractive
interaction, perhaps due to hydrogen bonding, between PS headgroups. We
find that the charged bilayers repel as expected when interlamellar forces
are measured by osmotic stress and X-ray diffraction. Questions therefore
arise as to why the same repulsion does not occur between charged groups on
the same bilayer and why the charged lipids are stiffer against bending. It
is likely that strong ion binding is responsible. Indeed, we found a few
years ago that protons bind so strongly to the charged lipids that, at 10-4
Molar concentrations (pH 4), the membrane is wrenched from the lamellar form
into an inverted hexagonal structure. An even more intriguing observation is
that, when the same lipids are used as a scaffold for protein channels, the
exposure to protons shifts channel gating. We
have thus become aware of the organizing power of small ions to change
membrane protein behavior. Guided by the unexpectedly strong attraction
between charged phospholipids and simple ions, we have begun to examine Li
ions with phosphatidylserine bilayers; our effort builds on the earlier work
of other investigators whose research suggests that Li ions act like protons.
We are now asking whether Li can stress lipid assemblies and consequently
whether low levels of Li modify channel gating. Structural studies have
progressed during the year, including work at CHESS (Cornell High Energy
Synchrotron Source), where we obtain high-resolution data needed for
measurement of fluctuations, forces, and stress in protein-hosting lipids. Harries D, Ben-Shaul A, Szleifer I. Enveloping of charged
proteins by lipid bilayers. J Phys Chem B 2004;108:1491-1496. Petrache HI, Tristram-Nagle S, Gawrisch K, Harries D, Parsegian
VA, Nagle JF. Structure and fluctuations of charged phosphatidylserine
bilayers in the absence of salt. Biophys J 2004;86:1574-1586. Sachs JN, Nanda H, Petrache HI, Woolf TB. Changes in
phosphatidylcholine headgroup tilt and water order induced by monovalent
salt: molecular dynamics simulations. Biophys J
2004;86:3772-3782. Sachs JN, Petrache HI, Woolf TB. Interpretation of small angle
X-ray measurements guided by molecular dynamics simulations of lipid
bilayers. Chem Phys Lipids 2004;126:211-223. Sachs JN, Petrache HI, Zuckerman DM, Woolf TB. Molecular
dynamics simulations of ionic concentration gradients across model membranes.
J Chem Phys 2003;118:1957-1969. van
der Waals forces Petrache, Podgornik,
Parsegian; in collaboration with French, Nagle Weak
compared with covalent bonds and electrical-charge interactions, charge-fluctuation
or van der Waals forces are not usually appreciated for their work among
membranes and macromolecules. The dominant force that coheres membranes and
proteins and source of the powerful surface tension at membrane interfaces,
van der Waals forces are again the dominant, perhaps sole attraction that
creates membrane multilayers or allows membranes to adhere to artificial
surfaces. This past year proved particularly productive as we learned to
formulate, measure, and modify these neglected interactions. The key was to
begin with the elements of physical theory and to relate the polarizability
of materials to the fluctuations of charges within them. We were thus able to
design experiments demonstrating how macromolecular organization responds to
deliberate changes in solution properties. We
were able to see how the addition of even the simplest salts to water around
lipid bilayers changes how membranes assemble into multilayers. For example,
we can see how the attraction between membranes varies when salts of
different ions, e.g., chloride compared with bromide, are dissolved in the
intervening water. Membrane multilayers swell by 50 percent with bromide but
not with chloride. We reformulated van der Waals forces between membranes to
show how they would respond to changes in solutions so as to have a strategy
to control membrane assembly. One unexpected byproduct was a collaboration
with engineers who have been using our equations to design production
procedures for thin-film resistors in computer chips. We expect the
collaboration to work to our benefit by providing us with experimental data
for use in computing van der Waals forces. Another unexpected result relates
to what was known about the friction between an ion and the water through which
it moves. Just as a charge-water interaction creates a drag, there is a
charge-fluctuation source of drag on a moving particle. Mkrtchian VM, Parsegian VA, Podgornik R, Saslow WM. Universal
thermal radiation drag on neutral objects. Phys Rev Lett 2003;91:220801.
Podgornik R, Podgornik R, Cosolutes
in molecular folding and association Harries, Parsegian; in
collaboration with Rau Driven
by the conditions set by smaller solutes, proteins “fold” and
“unfold.” Experimentally, such conditions are stated as intensive
variables (pH and other chemical potentials) as though small solutes were
infinite resources that come at an independently varied price.
Computationally, the finite spaces of simulation allow only small, fixed
numbers of these solutes. Combining the analytic Gibbs adsorption isotherm
with the computational To
monitor how small adherent molecules affect molecular association, we
measured the changes in binding free energy versus changes in water activity
for the specific binding of cyclodextrin with an adamantane derivative. In
the context of different salts and neutral agents, the dependence of the
binding constant on osmotic pressure suggests a release of 15 to 25 water
molecules from the interacting surfaces upon association, depending on type
of solute used. The observed dependence of binding free energy and enthalpy
with added solute indicates that the osmolytes interact primarily
enthalpically with the surfaces. Harries D, Podgornik R, Harries D, Strey HH, Sachs JN, Petrache HI, Zuckerman DM, Woolf TB. Molecular
dynamics simulations of ionic concentration gradients across model bilayers. J
Chem Phys 2003;118:1957-1969. COLLABORATORS Joel Cohen, PhD, University of the Pacific,
Monique Dubois, PhD, Service de Chimie
Moléculaire, CEA Saclay, Roger French, PhD, University of Pennsylvania,
Phildelphia, PA and Dupont Research Laboratories, Wilmington, DE Klaus Gawrisch, PhD, Laboratory of Membrane
Biochemistry and Biophysics, NIAAA, Per Lyngs Hansen, PhD, Vanik Mkrtchian, PhD, John Nagle, PhD, Donald Rau, PhD, Laboratory of Physical and
Structural Biology, NICHD, Thomas Zemb, PhD, Service de Chimie
Moléculaire, CEA Saclay,
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