The objective is to understand the structural basis for calcium regulation in cellular processes. It is proposed to study the structures of calcium regulating proteins in different functional states, principally using small angle solution scattering and spectroscopic techniques such as Fourier transform infra-red spectroscopy (FTIR) and circular dichroism (CD). Specifically, the work will focus on the class of structurally related, though functionally diverse, regulatory proteins that includes calmodulin, troponin C and parvalbumin. Initial studies have concerned the effects of the solution environment on the calmodulin and troponin C structures. Future work will be directed more toward the interactions of these proteins with other components in their biochemical systems. In particular, we wish to: o further study the influence of the solution environment on the structures of troponin C and calmodulin; o investigate the structural and functional consequences of covalent modifications of calmodulin; o study the interactions of extrinsic and intrinsic calmodulin with the subunits of phosphorylase kinase; o study the interactions of calmodulin with the calmodulin binding domain of myosin light chain kinase and related peptides. Small angle solution scattering will be used to characterize the overall shapes and interactions of components of these systems in different states of activation. FTIR and CD spectroscopy will be used to evaluate secondary structure content, and, in particular, to monitor changes in secondary structure that accompany changes in functional state. Interpretation of the low resolution structural information that is obtained using these techniques will be greatly enhanced by the availability of high resolution crystallographic data on the proteins. The long term goal is to understand the molecular mechanism for calcium regulation.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM040528-04
Application #
3298159
Study Section
Biophysical Chemistry Study Section (BBCB)
Project Start
1988-07-01
Project End
1993-06-30
Budget Start
1991-07-01
Budget End
1992-06-30
Support Year
4
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Los Alamos National Lab
Department
Type
Organized Research Units
DUNS #
City
Los Alamos
State
NM
Country
United States
Zip Code
87545
Heller, William T; Krueger, Joanna K; Trewhella, Jill (2003) Further insights into calmodulin-myosin light chain kinase interaction from solution scattering and shape restoration. Biochemistry 42:10579-88
Heller, William T; Finley, Natosha L; Dong, Wen-Ji et al. (2003) Small-angle neutron scattering with contrast variation reveals spatial relationships between the three subunits in the ternary cardiac troponin complex and the effects of troponin I phosphorylation. Biochemistry 42:7790-800
Heller, William T; Abusamhadneh, Ekram; Finley, Natosha et al. (2002) The solution structure of a cardiac troponin C-troponin I-troponin T complex shows a somewhat compact troponin C interacting with an extended troponin I-troponin T component. Biochemistry 41:15654-63
Trewhella, Jill; Krueger, Joanna K (2002) Small-angle solution scattering reveals information on conformational dynamics in calcium-binding proteins and in their interactions with regulatory targets. Methods Mol Biol 173:137-59
Krueger, J K; Gallagher, S C; Zhi, G et al. (2001) Activation of myosin light chain kinase requires translocation of bound calmodulin. J Biol Chem 276:4535-8
Vigil, D; Gallagher, S C; Trewhella, J et al. (2001) Functional dynamics of the hydrophobic cleft in the N-domain of calmodulin. Biophys J 80:2082-92
Gallagher, S C; Gao, Z H; Li, S et al. (2001) There is communication between all four Ca(2+)-bindings sites of calcineurin B. Biochemistry 40:12094-102
Krueger, J K; Gallagher, S C; Wang, C A et al. (2000) Calmodulin remains extended upon binding to smooth muscle caldesmon: a combined small-angle scattering and fourier transform infrared spectroscopy study. Biochemistry 39:3979-87
Wall, M E; Gallagher, S C; Trewhella, J (2000) Large-scale shape changes in proteins and macromolecular complexes. Annu Rev Phys Chem 51:355-80
Tung, C S; Wall, M E; Gallagher, S C et al. (2000) A model of troponin-I in complex with troponin-C using hybrid experimental data: the inhibitory region is a beta-hairpin. Protein Sci 9:1312-26

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