Understanding how a protein's sequence determines its three dimensional structure and, consequently, its function, is a fundamental problem of modern biology. Solving this problem becomes more urgent as an increasing number of human diseases and conditions are discovered to be attributable to misfolded proteins. These misfolded proteins include representatives of many structural and functional classes, one of which is the broad class of proteins possessing nucleotide-binding domains. Because these domains have highly conserved structures, an understanding of how they would be widely applicable. The enzyme dihydrofolate reductase (DHFR), which has an adenine-binding subdomain (ABD) similar to the classical nucleotide-binding motif, has been extensively characterized with respect to structure, catalytic mechanism and folding, making it an excellent system for investigating the folding properties of nucleotide-binding domains. The goals of this project are to modify DHFR in ways that facilitate the study of the thermodynamic stability and kinetic folding properties of its ABD by spectroscopic methods. Specifically, the cooperativity of the DHFR folding reaction will be reduced by the use of complementary fragments and circularly permuted variants, allowing characterization of the separate structural domains and their folding intermediates. Fragments with altered spectroscopic properties will be made in order to allow the formation of hydrophobic surfaces and tertiary contacts with the ABD to be monitored independently.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
5F32GM019401-02
Application #
2910037
Study Section
Molecular and Cellular Biophysics Study Section (BBCA)
Project Start
1998-05-01
Project End
Budget Start
1999-05-01
Budget End
2000-04-30
Support Year
2
Fiscal Year
1999
Total Cost
Indirect Cost
Name
Pennsylvania State University
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
City
University Park
State
PA
Country
United States
Zip Code
16802