The long-term goal of this research is to understand in terms of structure and dynamics the molecular mechanisms that regulate Syk and Src tyrosine kinase function in cellular signal transduction. A vast number of human diseases are linked to improper functioning of protein tyrosine kinases, and inhibitors of Src kinase are in clinical use. A trait of most signaling proteins is their engagement with multiple binding partners for the purpose of controlling cellular localization and enzymatic activity. This ability to form a variety of protein-protein interactions requires conformational flexibility, and thus characterization of this flexibility and diversity is a basic component of understanding, and eventually predicting, these molecular recognition events. To this end, solution NMR and computational methods will be used to investigate four aims related to conformational transitions and molecular recognition of Syk and Src-family protein tyrosine kinases in the context of B cell signaling. Syk association with other signaling proteins is regulated by tyrosine phosphorylation within the linker regions interdomain A (IA) and interdomain B (IB). (1) The hypothesis that the dissociation of Syk from the B cell receptor is triggered by structural disorder induced at a distance by phosphorylation of IA will be tested. (2) The structural basis for recognition of alternative patterns of phosphorylation of IB at two closely spaced tyrosines by various downstream effector proteins will be determined. Src kinases phosphorylate the B cell receptor which then forms the binding site for Syk to initiate intracellular signaling. (3) Activation of Src kinases is well known to require conformational change but the details of the transition pathway are unknown and will be investigated. (4) The receptor substrate interactions of Src kinase are will also be determined.

Public Health Relevance

The overall objective of this research is to define in structural and physical terms the molecular mechanisms that regulate protein tyrosine kinase (PTK) function. Because PTKs regulate signal transduction processes that are essential in all aspects of cell growth, differentiation, metabolism, and programmed cell death, abnormal regulation of PTKs is associated with numerous diseases. A trait of most protein kinases is that tyrosine phosphorylation controls their function, in particular their recognition for various cellular binding partners and their structural plasticity. The proposed research aims to elucidate mechanisms of this control for Syk and Src PTKs with the goal of contributing new knowledge that is useful in fighting human disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM039478-20
Application #
8245055
Study Section
Macromolecular Structure and Function B Study Section (MSFB)
Program Officer
Flicker, Paula F
Project Start
1991-08-01
Project End
2014-03-31
Budget Start
2012-04-01
Budget End
2013-03-31
Support Year
20
Fiscal Year
2012
Total Cost
$314,392
Indirect Cost
$103,230
Name
Purdue University
Department
Pharmacology
Type
Schools of Pharmacy
DUNS #
072051394
City
West Lafayette
State
IN
Country
United States
Zip Code
47907
Yu, Shuai; Huang, He; Iliuk, Anton et al. (2013) Syk inhibits the activity of protein kinase A by phosphorylating tyrosine 330 of the catalytic subunit. J Biol Chem 288:10870-81
Taylor, Gwen M; Ma, Lixin; Vogt, Volker M et al. (2010) NMR relaxation studies of an RNA-binding segment of the rous sarcoma virus gag polyprotein in free and bound states: a model for autoinhibition of assembly. Biochemistry 49:4006-17
Huang, He; Ozkirimli, Elif; Post, Carol Beth (2009) A Comparison of Three Perturbation Molecular Dynamics Methods for Modeling Conformational Transitions. J Chem Theory Comput 5:1301-1314
Zabell, Adam P R; Post, Carol Beth (2002) Intermolecular relaxation has little effect on intra-peptide exchange-transferred NOE intensities. J Biomol NMR 22:303-15
Zabell, Adam P R; Post, Carol Beth (2002) Docking multiple conformations of a flexible ligand into a protein binding site using NMR restraints. Proteins 46:295-307
Eisenmesser, E Z; Zabell, A P; Post, C B (2000) Accuracy of bound peptide structures determined by exchange transferred nuclear Overhauser data: a simulation study. J Biomol NMR 17:17-32
Gaul, B S; Harrison, M L; Geahlen, R L et al. (2000) Substrate recognition by the Lyn protein-tyrosine kinase. NMR structure of the immunoreceptor tyrosine-based activation motif signaling region of the B cell antigen receptor. J Biol Chem 275:16174-82
Eisenmesser, E Z; Post, C B (1998) Insights into tyrosine phosphorylation control of protein-protein association from the NMR structure of a band 3 peptide inhibitor bound to glyceraldehyde-3-phosphate dehydrogenase. Biochemistry 37:867-77