Research Interests

This laboratory probes the linkage between cooperative ligand binding, conformational change and enzyme activation by calmodulin, a regulatory calcium- binding protein. Calmodulin is the primary eukaryotic intracellular calcium receptor and serves as a second messenger to regulate cellular responses to transient calcium fluxes. It is clinically relevant for human physiology (for example, it is a target of anti-psychotic drugs) and is also found in plants and fungi. Cooperative binding of four calcium ions to calmodulin causes large conformational changes; these changes control the sites and extent of calmodulin activation of important cellular enzymes and structural proteins (see Figure).

In order to determine the states that are functionally significant in this complex network of interactions, it is necessary to develop and apply new methods to directly determine energetic and structural properties of calcium binding. Quantitative proteolytic footprinting and applications of multi-dimensional heteronuclear NMR are capable of monitoring individual residues or bonds during a titration representative of a cellular influx of calcium. These studies have shown that the two domains of calmodulin interact in different ways as calcium fills the four sites of the protein.

Calcium-dependent structural rearrangements of calmodulin also are monitored by changes in the fluorescence, circular dichroism and hydrodynamic properties. These combined approaches are aimed at elucidating molecular mechanisms of cooperativity in calmodulin by determining the differences in intrinsic binding affinity at the four calcium-binding sites of calmodulin and the extent and nature of inter- and intra-domain cooperativity. To dissect these interactions, we are studying the isolated domains of calmodulin and many mutants shown to have phenotypic effects on complexes of calmodulin with its target enzymes.

Computational molecular modeling is used to visualize and calculate properties of these proteins and serves as a complement to the experimental studies of ligand- linked conformational change. The goal is to combine energetic and structural data to formulate models that will explain how synchronized changes in calcium levels modulate many diverse physiological processes.

Recent Publications

Feldkamp, M.D., Gakhar, L., Pandey, N. and Shea, M.A. (2015) Stuctural Insights into Molecular Specificity of Trifluoperazine Binding to Calcium-Saturated Calmodulin and the Role of Met144. Proteins: Structure, Function and Bioinformatics. PDB ID - 4RJD (http://dx.doi.org/10.1002/prot.24781)

Newman, R.A., Sorensen, B.R., Kilpatrick, A.M., Shea, M.A. (2014) Calcium-dependent energetics of calmodulin domain interactions with regulatory regions of the Ryanodine Receptor Type 1 (RyR1).
Biophys Chem. 193-194:35-49. [DOI: 10.1016/j.bpc.2014.07.004] (http://www.ncbi.nlm.nih.gov/pubmed/25145833)

Wang, X., Boyken, S.E., Hu, J., Xu, X., Rimer, R.P., Shea, M.A., Shaw, A.S., Andreotti, A.H., and Huang, Y.H. (2014) Calmodulin and PI(3,4,5)P3 cooperatively bind to the Itk pleckstrin homology domain to promote efficient calcium signaling and IL-17A production. Sci. Signal 7:337. [DOI: 10.1126/scisignal.2005147] (http://www.ncbi.nlm.nih.gov/pubmed/25097034)

Shea, M.A., Correia, J.J., Brenowitz, M.D. (2011) Introduction: Twenty Five Years of the Gibbs Conference on Biothermodynamics. Biophysical Chemistry 159:1-5. (http://dx.doi.org/10.1016/j.bpc.2011.07.002)

Evans, T.I.A., Hell, J.W., Shea, M.A. (2011) Thermodynamic Linkage between Calmodulin Domains Binding Calcium and Contiguous Sites in the C-Terminal Tail of Ca V1.2. Biophysical Chemistry 159:172-187. (http://dx.doi.org/10.1016/j.bpc.2011.06.007)

Feldkamp, M.D., Yu, L., Shea, M.A. (2011) Structural Determinants of Apo Calmodulin Binding to the IQ-Motif of the NaV1.2 Voltage-Dependent Sodium Channel. Structure 19:733-47.

O'Donnell, S.E., Yu, L., Fowler, C.A., Shea, M.A. (2011) Recognition of Beta-Calcineurin by the Domains of Calmodulin. Proteins: Structure, Function, Bioinformatics 79:765-786.

Feldkamp, M.D., O'Donnell, S.E., Yu, L., Shea, M.A. (2010) Allosteric Effects of the Anti-Psychotic Drug Trifluoperazine on the Energetics of Calcium Binding by the Calmodulin. Proteins: Structure, Function, Bioinformatics 78: 2265-2282 (http://dx.doi.org/10.1002/prot.22739)

O'Donnell, S. E., Newman, R.A.., Witt, T.J., Hultman, R., Froehlig, J., Christensen, A.P., Shea, M.A (2009) Thermodynamics and Conformational Change Governing Domain-Domain Interactions of Calmodulin. in Methods in Enzymology 466: 503-526 Edited by Ackers, G.K., Holt, J., Johnson, M.L. (http://dx.doi.org/10.1016/S0076-6879(09)66021-3)

Evans, T.I.A. and Shea, M.A. (2009) Energetics of Calmodulin Domain Interactions with the Calmodulin Binding Domain of CaMKII. Proteins: Structure, Function, Bioinformatics 76:47-61. (http://dx.doi.org/10.1002/prot.22317)

O'Donnell, S. E., Newman, R.A., Witt, T.J., Hultman, R., Froehlig, J., Christensen, A.P., Shea, M.A. (2009) Thermodynamics and Conformational Change Governing Domain-Domain Interactions of Calmodulin in Methods in Enzymology. 466: 503-526 (Johnson, M.L., Holt, J., Ackers, G.K., editors)

Erickson, J.R., Joiner, M.L., Guan X., Kutschke, W., Yang, J., Oddis, C.V., Bartlett, R.K., Lowe, J.S., O'Donnell, S., Aykin-Burns, N., Zimmerman, M.C., Zimmerman, K., Ham, A.L., Weiss, R.M., Spitz, D.R., Shea, M.A., Colbran, R.J., Mohler, P.J., Anderson, M. (2008) A Dynamic Pathway for Calcium-Independent Activation of CaMKII by Methionine Oxidation. Cell 133: 462-474 (http://dx.doi.org/10.1016/j.cell.2008.02.048)

Theoharis, N.T., Sorensen, B.R., Shea, M.A. (2008) Neuronal Voltage-Dependent Sodium Channel Type II (NaV1.2) IQ Motif Lowers Calcium Affinity of the C-domain of Calmodulin. Biochemistry 47: 112-123 (http://dx.doi.org/10.1021/bi7013129)

Newman, R.A., VanScyoc, W.S., Jaren, O.R., and Shea, M.A. (2008) Domain-Specific Interactions of Calmodulin Bound to Melittin Preferentially Increases Clacium Affinity of Sites I and II. Proteins: Structure, Function, Bioinformatics 71:1792-812. (http://dx.doi.org/10.1002/prot.21861)

Akyol, Z., Gakhar, L., Sorensen, B.R., Hell, J.W., and Shea, M.A. (2007) The NMDA Receptor NR1 C1 Region Bound to Calmodulin: Structural Insights into Functional Differences between Homologous Domains. Structure 15: 1603-1617 (http://dx.doi.org/10.1016/j.str.2007.10.012)

Li, Q., Cooper, J.J., Altwerger, G.H., Feldkamp, M.D., Shea, M.A., and Price, D.P. (2007) HEXIM1 is a promiscuous double-stranded RNA-binding protein and interacts with RNAs in addition to 7SK in cultured cells. Nucleic Acids Research 35: 2503?2512. (http://dx.doi.org/10.1093/nar/gkm150)

Merrill, M.A., Malik, Z., Akyol, Z., Bartos, J.A., Leonard, A.S., Hudmon, A., Shea, M.A. and Hell, J.W. (2007) Displacement of a-Actinin from the NMDA Receptor NR1 C0 Domain by Ca2+/Calmodulin Promotes CaMKII Binding. Biochemistry 46:8485-97 (http://dx.doi.org/10.1021/bi0623025)

VanScyoc, W.S., Newman, R.A., Sorensen, B.R., Shea, M.A. (2006) Calcium Binding to Paramecium Calmodulin Mutants Having Domain?Specific Effects on Regulation of Ion Channels. Biochemistry 45: 14311-14324 (http://dx.doi.org/10.1021/bi061134d)

Ross J.B.A., Laws W., Shea M.A. (2006) Intrinsic Fluorescence in Protein Structure Analysis. in Protein Structures: Methods in Protein Structure and Structure Analysis: Luinescence Spectroscopy and Circular Dichroism, Eds. VN Uversky, EA Permyakov, Nova Science Publishers, Inc., New York. 55-72 ISBN 1600214045 (http://www.vanstockum.nl/product/7188809/Methods-In-Protein-Structure-And-Stability-Analysis.html)

Wang, B., Martin, S.R, Newman, R.A., Hamilton, S.L., Shea, M.A., Bayley, P.M., and Beckingham, K. (2004) Biochemical properties of V91G calmodulin: A calmodulin point mutation that deregulates muscle contraction in Drosophila. Protein Science 13: 3285-3297 (http://dx.doi.org/10.1110/ps.04928204)

Hines, R., Sorensen, B.R., Shea, M.A., Maury, W. (2004) PU.1 Binding to ets Motifs within the Equine Infectious Anemia Virus Long Terminal Repeat (LTR) Enhancer: Regulation of LTR Activity and Virus Replication in Macrophages. J. Virology 78: 3407?3418. (http://dx.doi.org/10.1128/JVI.78.7.3407-3418.2004)

Akyol, Z., Bartos, J., Jaren, O.R., Faga, L., Shea, M.A. and Hell, J.? (2004) Apo-Calmodulin Binds with its COOH-terminal Domain to the N-methyl-D-aspartate Receptor NR1 C0 Region. J. Biol. Chem. 279(3): 2166-2175. (http://dx.doi.org/10.1074/jbc.M302542200)

Faga, L.A., Sorensen, B.R., VanScyoc, W.S., and Shea, M.A. (2003) Basic Interdomain Boundary Residues in Calmodulin Decrease Calcium Affinity of Sites I and II by Stabilizing Helix-Helix Interactions. Proteins: Structure, Function & Genetics 50: 381-391. (http://dx.doi.org/10.1002/prot.10281)

Leonard, A.S., Bayer, K-U, Merrill, M., Shea, M.A., Schulman, H., and Hell, J.W. (2002) Regulation of CaMKII Docking to NMDA Receptors by Calcium/Calmodulin and a-Actinin. J. Biol. Chem. 277: 48441-48448 (http://dx.doi.org/10.1074/jbc.M205164200)

Jaren, O.R., Kranz, J.K., Sorensen, B.R., Wand, A.J., Shea, M.A. (2002) Calcium-Induced Conformational Switching of Paramecium Calmodulin: Changes in the Protein Backbone Observed by Heteronuclear NMR Studies. Biochemistry 41: 14158-14166. (http://dx.doi.org/10.1021/bi026340+)

VanScyoc, W.S., Sorensen, B.R., Rusinova, E., Laws, W., Ross, J.B.A., and Shea, M.A. (2002) Domain-Specific Fluorescence of Calmodulin: Phenylalanine Reports Exclusively on Calcium Binding to the N-Domain. Biophysical Journal 83: 2767-2790. (http://dx.doi.org/10.1529/biophysj.102)

Xiong, L-W, Newman, R.A., Rodney, G.C., Thomas, O., Zhang, J-Z, Persechini, A., Shea, M.A., and Hamilton, S.L. (2002) Lobe Dependent Regulation of Ryanodine Receptor Type One by Calmodulin. J. Biol. Chem. 277:40862-40870. (http://dx.doi.org/10.1074/jbc.M206763200)

Sorensen, B.R., Faga, L.A., Hultman, R., Shea, M.A. (2002) Interdomain Linker Increases Thermostability and Decreases Calcium Affinity of Calmodulin N-Domain. Biochemistry 41: 15-20. (http://dx.doi.org/10.1021/bi011718+)

Sun, H., Yin, D., Coffeen, L.A., Shea, M.A., and Squier, T.C. (2001) Mutation of Tyr138 Disrupts the Structural Coupling Between the Opposing Domains in Vertebrate Calmodulin. Biochemistry 40: 9605-9617. (http://dx.doi.org/10.1021/bi0104266)

VanScyoc, W. and Shea, M.A. (2001) Phenylalanine fluorescence studies of calcium binding to N-domain fragments of Paramecium calmodulin mutants show increased calcium affinity correlates with increased disorder. Protein Science 10:1758-1768. (http://dx.doi.org/10.1110/ps.11601)

Sorensen, B.R., Eppel, J.T., Shea, and M.A. (2001) Paramecium Calmodulin Mutants Defective in Ion Channel Regulation Associate with Melittin in the Absence of Calcium but Require it for Tertiary Collapse. Biochemistry 40:896-903. (http://dx.doi.org/10.1021/bi0023091)