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Description:
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Chemokines are important mediators of leukocyte migration . Chemokines bind
to G protein–coupled receptors (GPCR ) and cause conformational changes that trigger
intracellular signaling pathways involved in inflammation , injury healing , cancer ,
metastasis , and HIV infections . No direct structural information about any chemokine
receptor is available , but the structure of chemokines has been well studied . Structural
studies of chemokines coupled with cell -biological investigations may lead to a better
understanding of the mechanisms of chemokine -receptor interactions . In this Ph .D .
project , I studied the structural and functional relationship between chemokines and
chemokine receptors using NMR , X -ray crystallography , and mutagenesis approaches ,
coupled with several different cell -biology assays . We found that the conserved
“chemokine fold” can support different dimerization types in the chemokines family ,
although changing the dimers from CC - to CXC -type fold is not readily accomplished . I
also used an engineered covalently -bound dimer of the MIP -1β mutant , MIP -1β -A10C , to study the relationship between dimerization of chemokines and their interaction with
the CCR5 receptor . My results suggest that MIP -1β dimer neither bind nor activate the
CCR5 receptor . I also studied the biophysical properties of one N -terminal awkward
mutant of P2 -RANTES , which was originally selected by others from a phage display
using CCR5 -expressing cells . Although the NMR and X -ray crystal studies revealed that
the wild type RANTES is a tight homodimer , analytical ultracentrifugation reveals that
P2 -RANTES is a monomer in solution , the 1 .7 Å resolution X -ray crystal structure of
P2 -RANTES was found to be a packed tetramer . The mutated N -terminal residues play a
very important role in the tetramerization in the X -ray crystal structure . Finally I used
the HIV -1 env mediated cell -cell fusion assay to study the combination of chemokines or
chemokine variants with anti -HIV peptides C37 or /and T -20 . A surprisingly synergistic
effect was found between P2 -RANTES and C37 or T -20 . This combination stratagem
may lead to further useful drug combinations or drug delivery for more potent anti -HIV
treatments . |