Jeffrey W. Kelly
Research Interests
The central theme of our research is to
understand the chemistry and biology of peptides and proteins and
to develop new approaches for manipulating these properties with
purposefully-designed small molecules. We employ spectroscopic
and biophysical methods in combination with chemical synthesis
and recombinant DNA technology to accomplish these aims. Specific
research projects include:
(1) SPECTROSCOPIC AND BIOPHYSICAL STUDIES ON THE CONVERSION OF A
NORMALLY SOLUBLE AND FUNCTIONAL HUMAN PROTEIN INTO AMYLOID
FIBRILS. An invariant feature of the sixteen human amyloid
diseases (e.g., Alzheimer's Disease) is the presence of amyloid
fibrils proximal to dead or dying neurons. We are currently
studying three (3) amyloidogenic proteins, transthyretin,
gelsolin and A-beta, whose conversion from a well-defined
tertiary structure into a b-sheet rich quaternary structure
appears to be the causative agent in their associated diseases.
Analytical ultracentrifugation and mass spectrometry are employed
to study the quaternary structural changes while circular dichroism, fluorescence, and nuclear magnetic resonance
spectroscopic methods are utilized to study the tertiary
structural changes required for amyloid fibril formation.
Electron microscopy and atomic force microscopy are also employed
to study the mechanism of assembly of the amyloidogenic
intermediate. The dynamic characteristics of the amyloidogenic
intermediate are studied by H/D exchange employing electrospray
and MALDI MS. We now know that significant tertiary structural
changes are required for the conversion of these proteins into amyloid. The challenge remaining is to understand the details of
these changes, the dynamic properties of the intermediates, and
the self-assembly mechanism that yields amyloid.
(2) A CELL BIOLOGY APPROACH TO UNDERSTAND THE NEUROTOXICITY
EXHIBITED BY AMYLOID OR THE SOLUBLE AMYLOID PRECURSORS. Neurotoxicity, which is a common feature of all amyloid diseases,
is poorly understood. It is likely that amyloid exhibits toxicity
through multiple mechanisms. Using our expertise in the
preparation of amyloid and its soluble precusors in combination
with the cell and neurobiology expertise at Scripps, we have
embarked on a project to understand why amyloid fibrils kill
nerve cells.
(3) STRUCTURE-BASED SMALL MOLECULE APPROACH FOR PREVENTING THE
CONFORMATIONAL CHANGES ASSOCIATED WITH HUMAN AMYLOID DISEASE. We
have introduced a new therapeutic strategy for preventing amyloid
fibril formation whereby a high affinity ligand is used to
prevent the quaternary and tertiary or tertiary structural
changes required for fibril formation. In this project we employ
X-ray crystallography or NMR in combination with synthetic
chemistry to identify high affinity inhibitors. Recent cocrystal
structures of the normally folded form of transthyretin with
second generation inhibitors suggest new molecules to synthesize
and test which should be much improved. Our philosphy on
synthesis is to keep it simple, such that parallel syntheses can
be used to prepare libraries to quickly sort out structure
activity relationships.
(4) THE USE OF PEPTIDES, PEPTIDOMIMETICS, AND STRUCTURALLY DEFINED PROTEINS TO BETTER UNDERSTAND b-SHEET STRUCTURE IN AQUEOUS SOLUTION. Peptides < 40 amino acids in length typically do not adopt well-defined structures in aqueous solution, making it difficult to study beta-sheet tertiary structure with small peptides. In an effort to better understand b-sheet structure in aqueous solution, we have designed and synthesized unnatural amino acids which nucleate b-sheet formation when incorporated into a suprising range of sequences composed of six (6) or more natural amino acids. The challenge is to convert these dynamic beta-sheet structures into well-defined b-sheets that are protein like. In parallel we are studying the WW domainn (57 residues), which is an important and recently discovered signal transduction domain that adopts an isolated three standed b-sheet structure in aqueous solution as discerned from NMR and X-ray crystallography. This is an excellent protein to understand the contributions of H-bonding and hydrophobic interactions to beta-sheet structure. This will be accomplished by incorporating unnatural amino acids into the WW sequence by solid phase peptide synthesis to test the physical and spectroscopic properties of the resulting analogs.
(5) THE DEVELOPMENT OF COMPOSITE ORGANIC-INORGANIC SURFACES
EMPLOYING SELF-ASSEMBLING PEPTIDOMIMETICS THAT INDUCE INORGANIC
CRYSTAL GROWTH ON THEIR SURFACE. The use of small peptidomimetic
beta-sheet structures described above in combination with
electrostatic intermolecular interactions leads to b-sheet
building blocks which self-assemble to form a monolayer coating
at an air-water interface or on a hydrophobic surface. Controlled
self-assembly allows a periodic presentation of functional groups
on a surface which can nucleate mineral crystallization if the
lattice match is good. Taking clues from biology, we have
accomplished calcite mineralization. The future challenge and
potential of this approach is the formation of unatural inorganic
surfaces which can be used for a variety of applications such as
catalysis.
(6) DEVELOPING THERAPEUTIC STRATEGIES FOR
TREATING HUNTINGTON'S DISEASE. The main focus of this research is
to develop a therapeutic strategy for Huntington's Disease. We
aim to discover inhibitors that interfere with poly-Gln
beta-sheet mediated aggregation in the nucleus. Aggregation
inhibitors will be discovered through a selection scheme and a
screening approach. The selection utilizes an oligo-Gln sequence
fused to a protein that is critical for cell viability. When the oligo-(Gln)>39 portion of the fusion mediates aggregation, the
cells are unable to grow. Cell growth can be rescued by small
molecules that interfere with (Gln)>39 assembly. The
feasibility of a second approach, using small molecules to target
the mRNA of the expanded huntingtin protein to prevent its
translation, will also be evaluated.
(7) SYNTHESIS OF NATURAL AND UNNATURAL PRODUCTS LACKING AMIDE
BONDS FROM OLIGOPEPTIDES. Peptide synthesis has evolved to a
point where nearly any sequence can be prepared in very high
yield. We will take advantage of the stereochemical purity of
peptides and their capability to undergo intramolecular reactions
to prepare novel heterocycles that can bind to proteins for the
purpose of inhibiting protein-protein interactions. This work
will commence with oligo-Gln sequences.
Analogously, we will prepare the following sulfur-based heterocycles.:
Knowing the products we desire, we can utilize
mass spectrometry to identify productive reactions and in the
process we will likely
discover new types of chemical reactivity. Peptides have been
very much underutilized as starting materials in organic
synthesis and we
hope to change this.
(8) ATROPISOMERICALLY PURE BIARYLS AS LIGANDS FOR INHIBITING
PROTEIN-PROTEIN INTERACTIONS. Atropisomerization refers to
isomerization about the aryl-aryl bond of biphenyls substituted
with
X and Y substitutents where X does not equal Y. Depending on the size of X and Y it may be possible to resolve the R and S enantiomers from one another. We envision controlling the atropisomerization to make chiral ligands designed to target proteins by making X large and chiral, such that the reaction can only yield one atropisomer. One approach is to utilize:
The main goal of this project is to decipher the appropriate
methodology to do hindered aryl couplings such that the products
are
atropisomerically pure or nearly pure. These ligands should prove
to be excellent protein binders.