Regulated inter- and intramolecular protein interactions play central roles in signaling events that control lymphocytes responses. The Itk protein tyrosine kinase, a member of the Tec family, plays an important role in T cell development and activation. Previous work from this investigators postdoctoral studies revealed a novel intramolecular interaction involving proline residues and the SH3 domain in Itk. Here, the investigator proposes to extend those studies, based on preliminary data, that suggest additional regulatory intramolecular interactions which depend upon the TH and SH2 domain of Itk. High resolution multidimensional nuclear magnetic resonance (NMR) techniques will be used to explore this intramolecular interaction. Structures and interactions of protein fragments lacking the catalytic domain of the N-terminus of Itk will be studied to investigate the intramolecular associations that might regulate substrate binding and catalytic function in the intact protein. These studies may provide new knowledge regarding signal transduction by T and B cell antigen receptors. Furthermore, these studies may broaden the application of multi- dimensional NMR techniques to the study of larger multi-domain protein systems. Novel techniques combining recombinant protein expression and chemical ligation to generate proteins in which a single domain is isotopically labeled are proposed. This technology will not only provide detailed information about intramolecular interactions in Itk, but will be broadly applicable to the study of intramolecular interactions in other protein with complex domain structure.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI043957-05
Application #
6626347
Study Section
Allergy and Immunology Study Section (ALY)
Program Officer
Winter, David B
Project Start
1999-01-15
Project End
2003-12-31
Budget Start
2003-01-01
Budget End
2003-12-31
Support Year
5
Fiscal Year
2003
Total Cost
$146,897
Indirect Cost
Name
Iowa State University
Department
Biochemistry
Type
Other Domestic Higher Education
DUNS #
005309844
City
Ames
State
IA
Country
United States
Zip Code
50011
Devkota, Sujan; Joseph, Raji E; Boyken, Scott E et al. (2017) An Autoinhibitory Role for the Pleckstrin Homology Domain of Interleukin-2-Inducible Tyrosine Kinase and Its Interplay with Canonical Phospholipid Recognition. Biochemistry 56:2938-2949
Joseph, Raji E; Wales, Thomas E; Fulton, D Bruce et al. (2017) Achieving a Graded Immune Response: BTK Adopts a Range of Active/Inactive Conformations Dictated by Multiple Interdomain Contacts. Structure 25:1481-1494.e4
Roberts, Justin M; Tarafdar, Sreya; Joseph, Raji E et al. (2016) Dynamics of the Tec-family tyrosine kinase SH3 domains. Protein Sci 25:852-64
Chopra, Nikita; Wales, Thomas E; Joseph, Raji E et al. (2016) Dynamic Allostery Mediated by a Conserved Tryptophan in the Tec Family Kinases. PLoS Comput Biol 12:e1004826
Xie, Qian; Fulton, D Bruce; Andreotti, Amy H (2015) A selective NMR probe to monitor the conformational transition from inactive to active kinase. ACS Chem Biol 10:262-8
Devkota, Sujan; Joseph, Raji E; Min, Lie et al. (2015) Scaffold Protein SLP-76 Primes PLC?1 for Activation by ITK-Mediated Phosphorylation. J Mol Biol 427:2734-47
Boyken, Scott E; Chopra, Nikita; Xie, Qian et al. (2014) A conserved isoleucine maintains the inactive state of Bruton's tyrosine kinase. J Mol Biol 426:3656-69
Wang, Xinxin; Boyken, Scott E; Hu, Jiancheng et al. (2014) Calmodulin and PI(3,4,5)P? cooperatively bind to the Itk pleckstrin homology domain to promote efficient calcium signaling and IL-17A production. Sci Signal 7:ra74
Xie, Qian; Joseph, Raji E; Fulton, D Bruce et al. (2013) Substrate recognition of PLC?1 via a specific docking surface on Itk. J Mol Biol 425:683-96
Joseph, Raji E; Kleino, Iivari; Wales, Thomas E et al. (2013) Activation loop dynamics determine the different catalytic efficiencies of B cell- and T cell-specific tec kinases. Sci Signal 6:ra76

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