The goal of this proposal is to investigate the role of dynamics in the long-range coupling of allosteric sites in proteins. To accomplish this, site-directed mutagenesis will be used in combination with solution Nuclear Magnetic Resonance (NMR) relaxation techniques. Two protein systems are chosen for the proposed study. The first system, calmodulin, is a signaling protein that shows allosteric coupling between its calcium- and target-binding sites. Mutations in the vicinity of either binding site, known to affect calcium- or target-binding properties of calmodulin, will be introduced. The second system is drug-resistant mutants of HIV-1 protease, for which mutations distant from the active site were shown to be responsible for the impaired binding affinity towards protease inhibitors. Information about the backbone and sidechain dynamics of the mutant protein complexes with appropriate ligands will be obtained using NMR relaxation techniques. Dynamically perturbed residues will be identified by comparison with the wild-type protein complexes. The findings of this proposal will help to uncover the mechansim of allosteric coupling in monomeric proteins and the role of protein flexibility in the development of drug resistance.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
1F32GM071133-01
Application #
6790308
Study Section
Special Emphasis Panel (ZRG1-F04 (20))
Program Officer
Basavappa, Ravi
Project Start
2004-04-01
Project End
2007-03-31
Budget Start
2004-04-01
Budget End
2005-03-31
Support Year
1
Fiscal Year
2004
Total Cost
$42,976
Indirect Cost
Name
University of Pennsylvania
Department
Biochemistry
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Igumenova, Tatyana I; Lee, Andrew L; Wand, A Joshua (2005) Backbone and side chain dynamics of mutant calmodulin-peptide complexes. Biochemistry 44:12627-39