| Our research combines in vitro spectroscopic
and biophysical techniques, protein design and engineering
for the development of novel probes, and cellular imaging
studies to elucidate the mechanism of cellular signaling pathways.
We are focused on understanding pathways involved in metal/ion
homeostasis and apoptosis as they contribute to disease (Alzheimer’s
disease and cancer) or pathogenic processes (neurodegeneration).
Our lab uses a variety of techniques (molecular evolution,
phage display, rational protein design, and peptide synthesis)
to develop sensors and reporters based on fluorescence resonance
energy transfer (FRET). The sensors make use of a wide array
of fluorescent proteins that undergo energy transfer when
in close proximity and are designed such that binding of the
target causes a change in FRET that can be monitored with
a fluorometer (in vitro) or with a fluorescent microscope
(in cells). Because the sensors can be genetically encoded,
they can be targeted to specific subcellular locations, enabling
us to examine the spatial variability and compartmentalization
of different signaling processes within the context of live
cells. The advantage of such sensors is that they permit the
study of reactions while preserving the complex network of
interactions that occurs inside a cell and preserve the temporal
control/dynamics of different processes. In addition to visualizing
cellular processes, we are developing peptide-based probes
that permit acute perturbation of reaction pathways to enhance
our understanding of critical signaling reactions.
A powerful complement to these cellular studies are spectroscopic
and biophysical methods (absorption, circular dichroism, electron
paramagnetic resonance, fluorescence, fluorescence anisotropy,
stopped-flow, and surface plasmon resonance) that provide
detailed information on the bonding nature, kinetics, and
thermodynamics of protein-metal, protein-small molecule, and
protein-protein interactions.
Specific projects in the lab include 1) the development
of genetically encoded zinc sensors and examination of the
role of zinc in neuronal signal transduction, 2) the design
of peptide-based tools to perturb protein-protein interactions,
and 3) development of probes to study the connection between
amyloid-beta and calcium dysregulation.
Selected Publications:
Palmer, A. E., Jin, C, Reed, J. C., Tsien, R. Y., Bcl-2 mediated
alterations in endoplasmic reticulum Ca2+ analyzed with an
improved genetically encoded fluorescent sensor, Proc. Natl.
Acad. Sci., 2004, 101:50, 17404-17409 * This work was highlighted
in the following journals: BioTechniques, 2005, 38:1, p17;
Nature Reviews: Molecular Cell Biology, 2005, 6, p92
Shaner, N.C., Campbell, R.E., Steinbach, P.A., Giepmans, B.N.G.,
Palmer, A.E., Tsien, R.Y. Improved monomeric red, orange,
and yellow fluorescent proteins derived from Discoma red fluorescent
protein, Nature Biotechnology, 2004, 22, 1567-1572
Palmer, A.E., Szilagyi, R.K., Cherry, J.R., Jones, A., Xu,
F., Solomon, E.I. Spectroscopic characterization of the Leu513His
variant of fungal laccase: effect of increased axial ligand
interaction on the geometric and electronic structure of the
Type 1 Cu site Inorg. Chem., 2003, 42, 4006-4017
Campbell, R.E., Tour, O., Palmer, A.E., Steinbach, P.A., Baird,
G. S., Zacharias, D.A., Tsien, R.Y. A Monomeric Red Fluorescent
Protein Proc. Natl. Acad. Sci., 2002, 99, 7877-7882
Palmer, A.E., Quintanar, L., Severance, S., Wang, T.-P., Kosman,
D. J., and Solomon, E.I. Spectroscopic characterization and
O2 reactivity of the trinuclear Cu cluster of mutants of the
multicopper oxidase Fet3p, Biochemistry, 2002, 41, 6438-6448
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