The long-term objective of this proposed work is to establish a framework for using anti-peptide antibodies as probes of protein folding and dynamics. It is necessary first to investigate the mechanism of cross-reaction between intact proteins and anti-peptide antibodies. It is also necessary to develop a convenient method for studying the structures of proteins while they are bound to anti-peptide antibodies. We will study bovine pancreatic trypsin inhibitor (BPTI), a protein that has been well-characterized by biophysical and structural methods. Difference circular dichroism (CD) spectroscopy will be used to investigate the mechanism of cross-reaction at low resolution. The kinetics and thermodynamics of cross-reaction will be characterized. We will also determine if there is a correlation between the ability of peptides to fold, as judged by CD and nuclear magnetic resonance (NMR), and the ability of peptides to elicit cross-reacting antibodies. Amide proton exchange, combined with two-dimensional NMR spectroscopy (2D-NMR) will be used to study the structure of BPTI, while it is bound to anti-peptide antibodies. Hydrogen/deuterium exchange in the BPTI-antibody complex will be allowed to occur. Then, BPTI will be separated from antibody, and 2D-NMR will be used to quantitate the extent of exchange for individual amide protons in BPTI.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM037241-02
Application #
3292457
Study Section
Biophysics and Biophysical Chemistry A Study Section (BBCA)
Project Start
1986-07-01
Project End
1989-06-30
Budget Start
1987-07-01
Budget End
1988-06-30
Support Year
2
Fiscal Year
1987
Total Cost
Indirect Cost
Name
Whitehead Institute for Biomedical Research
Department
Type
DUNS #
076580745
City
Cambridge
State
MA
Country
United States
Zip Code
02142