Journal article
Interdomain cooperativity of calmodulin bound to melittin preferentially increases calcium affinity of sites I and II
Proteins, structure, function, and bioinformatics, Vol.71(4), pp.1792-1812
06/2008
DOI: 10.1002/prot.21861
PMID: 18175310
Abstract
Calmodulin (CaM) is the primary transducer of calcium fluxes in eukaryotic cells. Its two domains allosterically regulate myriad target proteins through calcium-linked association and conformational change. Many of these proteins have a basic amphipathic alpha-helix (BAA) motif that binds one or both CaM domains. Previously, we demonstrated domain-specific binding of melittin, a model BAA peptide, to Paramecium CaM (PCaM): C-domain mutations altered the interaction with melittin, whereas N-domain mutations had no discernable effect. Here, we report on the use of fluorescence and NMR spectroscopy to measure the domain-specific association of melittin with calcium-saturated ((Ca(2+))(4)-PCaM) or calcium-depleted (apo) PCaM, which has enabled us to determine the free energies of calcium binding to the PCaM-melittin complex, and to estimate interdomain cooperativity. Under apo conditions, melittin associated with each PCaM domain fragment (PCaM(1-80) and PCaM(76-148)), as well as with the C-domain of full-length PCaM (PCaM(1-148)). In the presence of calcium, all of these interactions were again observed, in addition to which an association with the N-domain of (Ca(2+))(4)-PCaM(1-148) occurred. This new association was made possible by the fact that melittin changed the calcium-binding preferences for the domains from sequential (C > N) to concomitant, decreasing the median ligand activity of calcium toward the N-domain 10-fold more than that observed for the C-domain. This selectivity may be explained by a free energy of cooperativity of -3 kcal/mol between the N- and C-domains. This study demonstrates multiple domain-selective differences in the interactions between melittin and PCaM. Our findings support a model that may apply more generally to ion channels that associate with the C-domain of CaM under low (resting) calcium conditions, but rearrange when calcium binding triggers an association of the N- domain with the channel.
Details
- Title: Subtitle
- Interdomain cooperativity of calmodulin bound to melittin preferentially increases calcium affinity of sites I and II
- Creators
- Rhonda A Newman - Department of Biochemistry, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, Iowa 52242-1109, USAWendy S Van ScyocBrenda R SorensenOlav R JarenMadeline A Shea
- Resource Type
- Journal article
- Publication Details
- Proteins, structure, function, and bioinformatics, Vol.71(4), pp.1792-1812
- Publisher
- United States
- DOI
- 10.1002/prot.21861
- PMID
- 18175310
- ISSN
- 0887-3585
- eISSN
- 1097-0134
- Grant note
- R01 GM 57001 / NIGMS NIH HHS T32 GM08365-13 / NIGMS NIH HHS
- Language
- English
- Date published
- 06/2008
- Academic Unit
- Molecular Physiology and Biophysics; Iowa Neuroscience Institute; Biochemistry and Molecular Biology
- Record Identifier
- 9984025299502771
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