Our laboratory focuses on using synthetic chemistry to construct macromolecules
of biological importance. We have developed a set of highly selective chemical
reactions that allow small peptide, nucleic acid or carbohydrate fragments
to be assembled selectively into functional macromolecules. For example, these
methods allow us to incorporate unnatural amino acids to probe fundamental
questions about protein folding, stability and enzymatic catalysis. In addition,
it is possible to generate proteins with specific fluorescent, PET or MRI imaging
agents, quantum dots or with crosslinking agents. Other studies are directed
towards HIV vaccine design and angiogenesis.
HIGHLIGHT
Dynamic Chemical Ligation
The ability to chemically modify biological macromolecules in a specific manner
underlies many of the methods and technologies used in modern research. This
specific tailoring of macromolecules has been enabled by the development
of highly chemoselective ligation (conjugation) chemistries that are characterized
by their chemoselectivity, reactivity and compatibility with neutral aqueous
buffers. However, the growing demands of research in the diverse areas of
analytical biochemistry, chemical biology, protein chemistry and nanotechnology
have pushed the limits of currently available ligation methods, especially
in terms of ligation kinetics. This year we demonstrated that the aromatic
amine, aniline is a potent nucleophilic catalyst for imine ligations that
form stable oximes and hydrazones from aldehyde and amine-labeled precursors.
We have used this catalyst to optimize imine reactions that achieve ligation
with rates over 1000 M-1 s-1, several orders of magnitude faster than currently
used ligation approaches. Such fast conjugation rates are essential if chemical
approaches are ever to compete with the rapid labeling possible using non-covalent
interactions such as biotin or antibodies. Importantly, since the ligation
rate is determined by the amount of catalyst, we can tune the reaction rate
to fit a desired application We currently are using this chemistry to efficiently
label proteinssuch as RANTES, albumin, myogolbin and annexin A5 with fluorescence,
PET and MRI imaging probes and to label nanoparticles. Future directions
of this research will focus its application in complex biological systems
such as cell surface and intracellular labeling strategies and the generation
of protein and carbohydrate arrays.
In addition to providing rapid and selective reaction rates, this catalytic approach
enables the reversible labeling of molecules. Using rapid reversible covalent
chemistry, we anticipate being able to select molecules out of dynamically exchanging
libraries based on binding affinity or target selectivity. We anticipate that
these reactions will enable us to develop reversible tagging strategies compatible
with complex biological systems. For example, a biotin could be removed from
a tagged probe while on a streptavidin column and rapidly replaced with a fluorophore
in a single step.
PUBLICATIONS 2006
1. Cremeens, M.E., Fujisaki, H., Zhang, Y., Zimmermann, J., Sagle, L.B., Matsuda,
S., Dawson, P.E. Straub, J.E., Romesberg, F.E. Efforts toward developing direct
probes of protein dynamics. J Am Chem Soc. 128(18):6028-9, 2006.
2. Sagle, L.B., Zimmermann, J., Matsuda, S., Dawson, P.E., Romesberg, F.E.
Redox-coupled dynamics and folding in cytochrome c. J Am Chem Soc. 128(24):7909-15,
2006.
3. Medintz, I.L., Clapp, A.R., Brunel, F.M., Tiefenbrunn, T., Uyeda, H.T.,
Chang, E.L., Deschamps, J.R., Dawson, P.E., Mattoussi, H. Proteolysis monitored
by FRET through quantum-dot-peptide conjugates. Nat Mater. 5(7):581-9, 2006.
4. Delehanty, J.B., Medintz, I.L., Pons, T., Brunel, F.M., Dawson, P.E., Mattoussi,
H. Self-assembled quantum dot-peptide bioconjugates for selective intracellular
delivery. Bioconjug Chem. 17(4):920-7, 2006.
5. Dirksen, A., Hackeng, T.M., Dawson, P.E. Nucleophilic Catalysis of Oxime
Ligation. Angew Chem. Int. Ed. Engl. 45: 7581-7584, 2006..
6. Sagle, L.B., Zimmermann, J., Dawson, P.E., Romesberg, F.E. Direct and high
resolution characterization of cytochrome C equilibrium folding. J. Am. Chem.
Soc. 128(44):14232-3, 2006.
7. Dirksen, A., Dirksen, S., Hackeng, T.M., Dawson, P.E. Nucleophilic catalysis
of hydrazone formation and transimination: implications for dynamic covalent
chemistry. J. Am. Chem. Soc. 128: 15602-15603, 2006.
8. Metanis, N., Keinan, E., Dawson, P.E. Synthetic Seleno-Glutaredoxin 3 Analogues
Are Highly Reducing Oxidoreductases with Enhanced Catalytic Efficiency. J.
Am. Chem. Soc. 128(51):16684-91, 2006.
9. Cardoso, R.M., Brunel, F.M., Ferguson, S., Zwick, M., Burton, D.R., Dawson,
P.E., Wilson, I.A. Structural Basis of Enhanced Binding of Extended and Helically
Constrained Peptide Epitopes. J. Mol. Biol. 2;365(5):1533-442006, 2007.
