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James M. Rini
Associate Professor & Graduate Coordinator
Ph.D., Toronto, 1986 |
Medical Sciences
Medical Sciences Building, Room 5360
416-978-0557
james.rini@utoronto.ca |
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Protein Structure and Molecular
Recognition
Research Synopsis
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Our research
interests are centred on the structural characterization
of protein-protein and protein-ligand complexes by x-ray
crystallography.
The specific research projects can be grouped into three
main categories:
i) protein-carbohydrate interactions and glycosyltransferases,
ii) adhesion molecules of the cadherin superfamily and
iii) cell surface signalling receptors.
Carbohydrate-Binding Proteins and Glycosyltransferases
Carbohydrate-binding proteins or lectins are a very diverse
class of proteins found in organisms ranging from bacteria
to man. Through specific interactions with various glycoconjugates
they are responsible for mediating processes including
pathogen-host recognition, immune surveillance, cell migration
and homing, protein folding and quality control in the
ER and the modulation of cell-matrix interactions. Through
x-ray diffraction analysis we are studying various lectin-saccharide
complexes as part of our efforts to determine the structural
basis for their functional properties. Further to our
interest in characterizing protein-carbohydrate interactions
we are studying a number of the glycosyltransferases responsible
for the biosynthesis of the N- and O-linked oligosaccharides
typical of secreted and cell surface glycoproteins. Our
recent x-ray crystal structure of N-Acetylglucosaminyltransferase
I has not only provided us with insight into the molecular
basis for specificity and catalysis, but through comparative
analysis it has allowed us to define a new protein superfamily.
This project is funded through the Protein Engineering
Network Centres of Excellence (PENCE) and the long-term
goal of this work is the development of anti-metastatic
and anti-inflammatory therapeutics.
Cadherins
The cadherins are a calcium dependent family of adhesion
molecules important in the formation and maintenance of
cell junctions. In man, epithelial cadherin (E-cadherin)
has been found to be a tumor invasion suppressor and is
thus an important determinant of the metastatic potential
of a given tumor cell. We have solved the x-ray crystal
structure of a two-domain fragment of E-cadherin in the
presence of calcium. The structure has defined the role
played by metal binding in this important class of calcium-dependent
adhesion molecules. Nevertheless, a complete description
of how the cadherins promote cell junctions, at the molecular
level, remains unresolved. Through further crystallographic
analysis, mutagenesis/cell transfection assays and binding
studies, we hope to shed light on this important process.
Cell Surface Signalling Receptors
Complement component C3 is central to host defense as
its proteolytic activation is the point of convergence
of the classical, alternative and lectin pathways of complement
activation. Furthermore, the antigen bound C3d(g) fragment
of C3 serves as ligand for complement receptor 2 (CR2)
on B cells during a normal antibody response. This receptor-mediated
interaction is of considerable interest as it provides
an important link between the innate and adaptive arms
of the immune system. Our x-ray crystal structure of C3d
has provided an understanding of the mechanistic basis
for its attachment to pathogenic cell surfaces, as well
as insight into the structural basis for its ability to
interact with CR2. Our current efforts on this project
are directed toward characterization of the CR2-C3d interaction.
We have recently solved the x-ray crystal structure of
Discoidin I, a D. discoideum lectin important in the cell-streaming
phase of this organism's life cycle. The discoidin homology
domain has now been found in a number of tyrosine-kinase
signalling receptors and cell surface molecules important
in neuronal pathfinding in mammals. Through analysis of
members of this family we hope to determine the functional
role played by the discoidin domain in these molecules.
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Selected Publications
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Chen, W., Ünligil,
U.M., Rini, J.M., and Stanley P. (2001) Independent Lec1A
CHO Glycosylation Mutants Arise from Point Mutations in
N-Acetylglucosaminyltransferase I that Reduce Affinity
for Both Substrates. Molecular Consequences Based on the
Crystal Structure of GlcNAc-T1. Biochemistry, 40, 8765-72.
Ünligil, U.M., Zhou, S., Yuwaraj, S., Sarkar, M., Schachter,
S. and Rini, J.M. (2000). X-ray Crystal Structure of Rabbit
N-Acetylglucosaminyltransferase I: Enzyme Mechanism and
a New Protein Superfamily. EMBO J., 19, 5269-5280.
Rini, J.M. and Sharon, N. (2000). Glycosyltransferases,
sugar nucleotide transporters and bacterial surface lectins
- at the cutting edge of glycobiology. Curr. Opin. Struct.
Biol. 10:507-509.
Ünligil, U.M. and Rini. J.M. (2000). Glycosyltransferase
structure and mechanism. Curr. Opin. Struct. Biol. 10:510-507.
Nagar, B., Jones, R.G., Diefenbach, R.J., Isenman, D.E.,
Rini, J.M. (1998). X-ray Crystal Structure of C3d: a C3
fragment and Ligand for Complement Receptor 2. Science,
280, 1277-1281.
Seetharaman, J., Kanigsberg, A., Slaaby, R., Leffler,
H., Barondes, S.H. and Rini, J.M. (1998). X-ray Crystal
Structure of the Human Galectin-3 Carbohydrate Recognition
Domain at 2.1 Å Resolution. J. Biol. Chem., 273, 13047-13052.
Nagar, B. Overduin, M. Ikura, M. and Rini, J.M. (1996).
Structural Basis of Calcium Induced E-Cadherin Rigidification
and Dimerization. Nature, 380, 360-364.
Rini, J.M.(1995). Lectin Structure. Ann. Rev. Biophys.
Biomolec. Struct., 24:551-557. |
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