Research
IRG IV
Bio-Interfacial Science
Faculty (* = coordinators): P. Cluzel, A. Dinner, R. Ismagilov, B. Kay*, S. Kent, S. Kron, K.Y. Lee, M. Mrksich*, D. Preuss, N. Scherer
This IRG addresses three important themes at the interface of the materials and biological sciences. One broad theme applies a materials science approach to developing biochips for quantitative characterization of biological activities. A second theme is developing materials and methods to understand the principles that underlie polyvalent interactions in biology — including the adhesion and migration of animal and plant cells — with the aim of translating these principles to materials research. The third, emerging, theme investigates biological approaches — based on the assembly of protein units — to prepare novel materials that are structurally ordered at the nanometer scale and incorporate dynamic properties. An overarching facet of this IRG is the vision that the methods and tools that are common to materials science, chemistry and physics offer new opportunities for understanding the bio/materials interface and, in turn, provide new opportunities for controlling the interface for a range of applications. The goals of this IRG are to foster and support close collaborations that are essential to pursue these themes, to gain a fundamental understanding of the bio/materials interface, and to realize practical materials and devices that combine biological and materials components.
Biochips. This effort has developed quantitative appraoches
to biochips—including arrays of peptide, protein, carbohydrate and
lipid—that can characterize protein binding and enzymatic activities.
This IRG applies an integrated approach, combining surface chemistry, biophysics,
theory, molecular and cell biology to engineer interfaces that give quantitative
information on biological activities. The effort started with the demonstration
by Mrksich and Kron of a solid-phase assay for quantitative analysis of
kinase activities. Ismagilov and Mrksich have extended this work to integrate
multi-analyte assays with a microfluidic platform to profile a panel of
kinase activities. Dinner and Mrksich are combining theory and experiment
to understand the role of ‘rebound’ in the kinetics of interfacial
enzyme reactions, and in one example have shown that recombinant proteins
containing multiple copies of the same enzyme domain catalyze an interfacial
reaction with the same rate as a mono-domain enzyme. To translate these
assays to more complex biomolecules, such as proteins and antibodies, Kent
and Mrksich have developed methods for immobilizing proteins with control
over the density and orientation. Kent has recently extended the native
chemical ligation reaction to non-Cys ligation sites for the total synthesis
of peptides and proteins containing Cys residues. Kay and Mrksich have developed
and applied surface chemistries for the immobilization of his-tagged proteins
and demonstrated the measurement of protein-protein interactions from cellular
lysates. The IRG has further emphasized the development of label-free detection
formats that may have a substantial impact on protein microarray technology.
The investigators have demonstrated a high throughput strategy for testing
protein-protein interactions among a set of recombinant proteins and have
performed several thousand assays in a bacterial proteome. Other work is
applying the label-free biochip strategies to characterize the enzyme activities
that underlie the histone code that provides a level of epigenetic control
to gene expression. This theme within the IRG has identified a broad set
of opportunities where the collaboration of physical and biological scientists
is yielding new materials for the development of bioanalytical systems.
Polyvalency in Biology and Materials. This theme of the
IRG applies experimental and theoretical techniques from materials science
to investigate the properties of polyvalent systems in cell biology. The
program has a two-fold goal: understanding the design rules for engineering
materials used in biological applications (e.g., scaffolds for tissue engineering)
and revealing design principles that can be applied to materials science.
Polyvalent systems are those wherein the behaviors of cells depend on the
presentation of many copies of a ligand, and do not follow directly from
an understanding of a single ligand-receptor interaction. The IRG combines
biology (Kron, Preuss), theory (Dinner) and physical methods (Cluzel, Ismagilov,
Mrksich) to address two problems: the movement of microbes towards nutrient
sources and the migration of adherent cells. Both systems rely on molecular
gradients of ligands, which direct how cells migrate, differentiate, and
organize into patterns. The IRG has developed methods for imposing chemotactic
(attraction or repulsion to a gradient of soluble molecules) and haptotatic
(attraction or repulsion to a gradient of immobilized molecules) gradients
on cells. Ismagilov has developed microfluidic approaches to generating
time-dependent gradients of multiple molecules and with Mrksich has patterned
an array of squares that each present a gradient of cell-adhesive ligands.
These substrates have been used to characterize the response of a cellular
cytoskeleton to a non-uniform density of adhesive ligands. In the second
model, Ismagilov and Preuss are continuing to develop a three-dimensional
microfluidic guidance assay to study the chemo-attraction of pollen tubes
in plants, using Arabidopsis thaliana as the model system. Formation of
attractant gradients in a three dimensional devices has been characterized
using both experiments and modeling. These devices are being used to determine
how the shape of the attractant gradient affects pollen tube guidance. The
three-dimensional system is also being used to identify the signaling molecules
in the exudates from live ovules involved in guidance of pollen tubes. Ismagilov’s
work on gradients also includes characterization of thermal gradients around
a living, developing embryo, carried out in the context of this program.
Laminar flow in microfluidics was used to create a temperature step around
a live Drosophila embryo, which enabled the study of the influence of temperature
on early patterning and nucleation of the embryo. The flow and temperature
profile around the embryo were characterized both experimentally and by
numerical simulations. Taken together, these projects are developing the
materials science for a range of applications in basic and applied biology,
and are revealing physical principles, with the support of models, to understand
the properties of polyvalent systems.
Bioinspired Nanostructured Materials. This theme seeks to develop
and exploit the assembly processes inherent to biology for the fabrication
of materials with unique properties. This paradigm is especially relevant
to materials with ordered structure at the 10-50 nm length scale, since
these structures are outside the scope of synthetic methods and are too
fine to be routinely prepared using lithographic methods. The strategy relies
on preparing protein building blocks which, under specified conditions,
will self-assemble into well-defined periodic structures. Mrksich has prepared
proteins that contain a calmodulin domain that undergoes conformational
changes in response to small molecule binding, and have created hydrogels
based on the cross-linking of this protein into a ethylene glycol-based
spacer. The resulting gels show macroscopic volume changes in response to
specific small molecule stimuli. To further characterize the dynamic motions,
Scherer is developing a unique interferometric fluorescence microscope that
will allow nanometer-scale determination of single molecule (or quantum
dots or metal nanoparticle scatterers) in discreet volumes. This work is
combined with single molecule biophysics and simulation studies to understand
the relationships between protein dynamics and macroscopic dynamics. Scherer
has utilized single molecule nonequilibrium force measurements to demonstrate
that the chemical adhesive in mussels, the tyrosine derivative 3,4-dihydroxy-L-phenylalanine
(dopa), creates a very large adhesion force to both titanium or organic
surfaces dependent upon its chemical oxidation state. Repeated force-extension
measurements allowed observing switching of the chemical state between the
dopa (strong adhesion to metal) and dopa–quinone (weaker adhesion)
forms. A second study has established the mechanistic underpinnings of anisotropic
stabilities of photoactive yellow protein by pulling along distinct axes
of individual polyprotein constructs. In ongoing research on the latter
topic, Dinner and Scherer employ a comprehensive analysis and simulation
approach to understand the mechanism of anisotropic protein stability; applying
a non-equilibrium statistical mechanical (i.e. Jarzynski equality) analysis
of the data and obtained atomistic insight into the mechanistic differences
via steered molecular dynamics simulations.
From annual report to the NSF, March 2007