Principal Investigator/Program Director (Last, first, middle): Wang, Ross, E.
Steric-free labeling strategies to study disease-related non-histone substrates of post-
translational modifications
The proposed research will interrogate fluorine-thiol based bioorthogonal reactions for steric-free labeling of
post-translational modifications (PTMs). Despite recent advances in biomedical research, diseases are still
haunting human beings. Many cancer types remain lethal and are resistant to traditional therapy, while most
inflammatory diseases result in chronic or long-term burdens, and there is a lack of selective immunosuppressive
on the market. Thus, new therapeutic approaches are needed. PTMs have recently emerged as a class of
important biological pathways that could unravel potentially novel therapeutic targets. PTMs modify existing
proteins with additional chemical functionalities to modulate protein function, and thereby mediate various cellular
activities. Dysregulation of PTM-related proteins has been reported to be key to certain human diseases such
as cancer and inflammatory disorders. Yet, the identity of these non-histone proteins has not been fully
elucidated. Current research in this area heavily relies on chemical proteomics, which tags target proteins with
alkyne or azide- modified PTM cofactors or precursors. These chemical tags on PTM sites can later be
derivatized in situ through bio-orthogonal `click chemistry' with a fluorophore for imaging or a biotin affinity probe
for pull down and proteomics-based target identification. However, these alkyne/azide- based tags are bulky in
length and size, and for many cases alkyne or azide- tagged PTM precursors/cofactors (e.g. for acetylation,
methylation) were barely incorporated onto substrates, thereby limiting PTM-related target identification.
To solve this issue, an innovative chemical tagging approach will be developed here, which could be steric free,
and can be broadly used for the global profiling of proteins related to various types of PTMs that are important
to the onset or relapse of human diseases. We hypothesize that unlike azides and alkynes, fluorine labeling can
best mimic the intrinsic carbon-hydrogen bond, and is thereby steric free and generally applicable to tag PTM
cofactors/precursors. The first research thrust seeks to utilize chemical synthesis to derivatize common PTM
cofactor/precursors with fluorine, then to hijack the PTM biological pathway to tag substrate proteins with fluorine,
and finally to modify the fluorinated substrate proteins with thiol-derivatized probes for imaging or protein
identification. The second research thrust involves the application of this fluorine-thiol tagging strategy to
interrogate the substrate proteins of PTMs, which are potentially important to the survival of cancer cells, but not
regular cells. The final thrust will investigate the substrates of PTMs which mediate human T cell activation, and
are potentially pivotal to the modulation of auto-reactive immune responses.
Completion of these research goals will result in the invention of a chemical labeling method that efficiently and
accurately identifies disease-associated subcellular components. This strategy will lead to a systemic dissection
of PTM-related disease signaling. The achieved results will greatly accelerate disease diagnosis and treatment,
and will also result in a tool box of steric-free PTM probes for the benefit of the scientific community.
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