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Boris Steipe
Associate Professor
Ph.D., Ludwig-Maximilians-Universität,
Munich, Germany 1990 |
Medical Sciences Building, Room 5368
416-946-7741
boris.steipe@utoronto.ca |
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Informational Protein Folding
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Research Synopsis
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PH-domain architecture:
The bipartite architecture of PH domains (variable loops
on structurally conserved framework) makes them well suited
for epitope display. This schematic shows how the variable
loops of the PH-domain (blue) align with the three major
antigen-binding loops of an immunoglobulin domain (green).
We propose that comparable structural variability can
be realized with the PH-domain, which is only half as
large as an Fv-fragment, is monomeric and requires no
structural disulfide bridge for folding. |
The methods
and topics of life science research have changed profoundly
in the last decade. Genomic science has shifted our attention
from hypothesis-driven studies of individual molecules
to phenomenological approaches that target comprehensive
views of the living cell and its dynamics. Proteomics
is setting its sights on a description of the entire complement
of cellular components and their interactions.
This development has been technology driven to a large
extent; as a corollary we see that the applications domain,
especially the biotechnology industry, contributes an
unusually large fraction of its momentum. We - as basic
scientists in the traditional centres of collaboration,
education and non-directed discovery - are challenged
not to fall behind: experience shows that sustained progress
in the private sector critically depends on the academic
sector to provide an uninterrupted stream of insight and
innovations in the long-term.
With this in mind, our program of biomolecular engineering
research emphasizes the investigation of engineering principles
and strategies that may contribute to an understanding
of biomolecular systems at a level that goes beyond phenomenological
descriptions and short-term application issues. Our topics
span bioinformatics, protein engineering and biomolecular
nanotechnology. At the core of the program is the analysis
and integration of genomics and proteomics data to guide
protein engineering, the experimental validation of our
predictions, and the application of results for molecular
medicine and bio-nanotechnology in a mid- to long-term
timeframe.
The cohesive element in our research projects ist the
quest to understand complexity in biomolecular systems.
Complexity arises from a context dependent behaviour of
system components and we observe complexity in many hierarchical
layers of structure formation and generation of function,
from the genome to the living cell. We focus our work
mainly on proteins since protein folding is the quintessential
paradigm of self-organising molecular systems. Based on
our concepts to address complexity, we develop strategies
and algorithms to analyse proteins and engineer them in
predictable ways. We apply our understanding to interesting
model proteins and biomolecular assemblies and we aim
to spin out successful concepts to biotechnology and medicine.
"Engineering" implies the rational application of well
understood principles with predicted outcome. Since structured
biomolecules are complex systems, the effects of changes
are difficult to predict. "Protein Engineering" thus may
be regarded to be an oxymoron, a contradiction in terms.
Nevertheless, we have been able to provide at least some
rationality to the engineering of antibody domains and
to the designed construction of biomolecular nanoassemblies.
We anticipate further progress in the application of our
strategies.
Layers of biological complexity: (a) Protein sequences are represented in the genome, but transcriptional regulation already provides the first unknown, as to which genes are being transcribed under which circumstances. As the transcript is further processed from left to right in this schematic, regulated alternative splicing will significantly change the m-RNA message of most genes. Finally, after translating the protein, the polypeptide folds to a strcuture that cannot be predicted from knowledge of the sequence information. Thus while cellular behavior as a result of protein function as a result of protein structure as a result of protein sequence is wholly represented in the genome, we have no way of assembling the latter from the former, unless the entire context is completely specified at the same time. This is a little like having to know the answer, in order to be able to ask a question. (b) The problem of hierarchical layers of complexity, where one layer cannot be defined before the entire underlying layer is specified, is complicated further when individual proteins assemble structurally into functional molecular machines, or functionally into metabolic or signalling pathways, and other higher-order cellular subsystems. Dynamic switching by posttranslational modifications becomes an issue, and protein sorting into different compartments must be taken into account, among many other aspects. (c) No less complex, and no more predictable from first principles, is the self-assembly of cells into organs and organisms during development.
Yet the entire process is self-organized, robust and reproducible.
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Selected Publications
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Wiehler J, von
Hummel J and Steipe B. (2001) Mutants of Discosoma red
fluorescent protein with a GFP-like chromophore FEBS Letters
487:384-389.
PDF Niggemann, M. and Steipe, B. (2000) Exploring Local
and Non-local Interactions for Protein Stability by Structural
Motif Engineering J Mol Biol 296: 181-195.
Ohage, E.C. and Steipe, B. (1999) Intrabody construction
and expression I: The critical role of VL domain stability
J Mol Biol 291: 1119-1128.
Ohage, E.C., Wirtz P., Barnikow, J. and Steipe, B. (1999)
Intrabody construction and expression II: A synthetic
catalytic Fv J Mol Biol 291: 1129-1134.
Wirtz P. and Steipe, B. (1999) Intrabody construction
and expression III: Engineering hyperstable VH domains
Protein Science 8: 2245-2250.
Ohage, E.C., Graml, W., Walter, M.M., Steinbacher, S.
and Steipe, B. (1997) beta-turn Propensities as Paradigms
for the Analysis of Structural Motifs to Engineer Protein
Stability Protein Science 6: 233-241.
Korndörfer, I., Steipe, B., Huber, R., Tomschy, A. and
Jaenicke, R. (1995) The crystal structure of holo-glyceraldehyde-3-phosphate
dehydrogenase from the hyperthermophilic bacterium Thermotoga
maritima at 2.5 Å resolution. J Mol Biol 246: 511-521.
Steinbacher, S., Seckler, R.,
Miller, S., Steipe, B., Huber,R. and Reinemer, P. (1994)
Crystal structure of P22 tailspike protein: interdigitated
subunits in a thermostable trimer Science 265: 383-386
Steipe, B., Schiller, B., Plückthun, A. and Steinbacher,
S. (1994) Sequence statistics reliably predict stabilizing
mutations in a protein domain. J Mol Biol 240: 188-192.
Steipe, B., Plückthun, A. and Huber, R. (1992) Refined
crystal structure of a recombinant immunoglobulin domain
and a complementarity-determining region 1-grafted mutant.
J Mol Biol 225: 739-753 |
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