Three-dimensional structure of calmodulin

Babu, Y.S.; Sack, John S.; Greenhough, Trevor J.; Bugg, Charles E.; Means, Anthony R.; Cook, William J.

doi:10.1038/315037a0

Cited by 1,049 publications

(645 citation statements)

References 42 publications

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“…Direct measurement of Ca 2ϩ binding to apo-CaM plus FFF confirms the enhancement of the N-domain Ca 2ϩ affinity. 2 For these M13-related peptides and for CBP1 the rates observed with T26W CaM are similar to, although somewhat faster than, the rates observed with Drosophila CaM. For the mastoparans, however, the rates observed with T26W CaM are significantly faster (Table II).…”

Section: Discussionmentioning

confidence: 69%

“…The affinities of the peptides for CaM were measured by direct fluorometric titration (10) or by competition with a peptide of known affinity. 2 Stopped-flow Measurements-Stopped-flow experiments were performed on a Hi-Tech SF61-MX stopped-flow spectrophotometer. Trp, Tyr, and Quin2 fluorescence signals were monitored using excitation wavelengths of 290, 280, and 334 nm, respectively, and emission cut-on filters of 320, 305, and 370 nm, respectively.…”

Section: Methodsmentioning

confidence: 99%

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Kinetic Control of the Dissociation Pathway of Calmodulin-Peptide Complexes

Brown¹,

Martin²,

Bayley³

1997

Journal of Biological Chemistry

105

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The mechanism of dissociation reactions induced by calcium chelators has been studied for complexes of Drosophila calmodulin with target peptides, including four derived from the skeletal muscle myosin light chain kinase target sequence. Reactions were monitored by fluorescence stopped-flow techniques using a variety of intrinsic probes and the indicator Quin2. For most of the complexes, apparently biphasic kinetics were observed in several fluorescence parameters. The absence of any obvious relationship between dissociation rates and peptide affinities implies kinetic control of the dissociation pathway. A general mechanism for calcium and peptide dissociation was formulated and used in numerical simulation of the experimental data. Unexpectedly, the rate of the slowest step decreases with increasing [peptide]/[calmodulin] ratio. Numerical simulation shows this step could contain a substantial contribution from a reversible relaxation process (involving the species Ca 2 -calmodulin-peptide), convolved with the following step (loss of C-terminal calcium ions).The results indicate the potentially key kinetic role of the partially calcium-saturated intermediate species.They show that subtle changes in the peptide sequence can have significant effects on both the dissociation rates and also the dissociation pathway. Both effects could contribute to the variety of regulatory behavior shown by calmodulin with different target enzymes.

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Section: Discussionmentioning

confidence: 69%

Section: Methodsmentioning

confidence: 99%

Kinetic Control of the Dissociation Pathway of Calmodulin-Peptide Complexes

Brown¹,

Martin²,

Bayley³

1997

Journal of Biological Chemistry

105

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“…The Ile-27 residue is one that participates in the hydrophobic cleft that is exposed upon Ca2+ binding (Babu et al, 1985(Babu et al, , 1988. If chemical shift changes of Ile-27 CyH, are indicative of formation of the hydrophobic cleft, then the results shown here suggest that the binding of a single calcium ion in either site in the N-terminal domain can activate the conformational "switch" that organizes the residues involved in forming the hydrophobic cleft.…”

Section: Conformational Effectsmentioning

confidence: 81%

“…The CaHs from Thr-26 and Asp-64 are positioned directly across from each other in a region of 0-sheet that lies between sites I and I1 (Kinemage 4; Babu et al, 1985Babu et al, , 1988Seeholzer & Wand, 1989). In wild-type CaM, these peaks titrate upfield almost simultaneously in fast exchange with midpoints of the change in chemical shift observed at 3.1 (Thr-26) and 3.0 (Asp-64) equivalents of Ca2+ (Fig.…”

Section: Site I and Ii Mutantsmentioning

confidence: 95%

A series of point mutations reveal interactions between the calcium‐binding sites of calmodulin

et al. 1992

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Calmodulin is a member of the "EF-hand" family of Ca'+-binding proteins. It consists of two homologous globular domains, each containing two helix-loop-helix Ca2+-binding sites. To examine the contribution of individual Ca2+-binding sites to the Ca2+-binding properties of CaM, a series of four site-directed mutants has been studied. In each, the glutamic acid at position 12 in one of the four Ca2+-binding loops has been changed to a glutamine. One-dimensional 'H-NMR has been used to monitor Ca2+-induced changes in the mutant proteins, and the spectral changes observed for each mutant have been compared to those for wild-type CaM. In this way, the effect of each mutation on both the mutated site and the other Ca2+-binding sites has been examined. The mutation of glutamate to glutamine at position 12 in any of the EF-hand Ca'+-binding loops greatly decreases the Ca2+-binding affinity at that site, yet differs in the overall effects on Ca2+ binding depending on which of the four sites is mutated. When the mutation is in site I, there is only a small decrease in the apparent Ca2+-binding affinity of site 11, and vice versa. Mutation in either site I11 or IV results in a large decrease in the apparent Ca2+-binding affinities of the partner C-terminal site. In both the N-and C-terminal domains, evidence for altered conformational effects in the partners of mutated sites is presented. In the C-terminus, the conformational consequences of mutating site 111 or site IV are strikingly different.

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“…CaM consists of two globular domains, each comprising two EF-hand motifs and a flexible linker [1,2]. A pair of Ca 2+ -binding sites is located on each globular domain.…”

Section: Introductionmentioning

confidence: 99%

Conformational dynamics of yeast calmodulin in the Ca²⁺‐bound state probed using NMR relaxation dispersion

et al. 2012

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a b s t r a c tMost calmodulin (CaM) in apo and Ca 2+ -bound states show a dumb-bell-like structure, involving the N-and C-terminal domains, connected with a flexible linker. However, Ca 2+ -bound yeast calmodulin (yCaM) takes on a unique globular structure; the target-binding site of this protein is autoinhibited. We applied NMR relaxation dispersion experiments to yCaM in the Ca 2+ -bound state. The amide 15 N and 1 H N relaxation dispersion profiles indicated the presence of conformational dynamics for specific residues at the interface between the N-and C-terminal domains. We conclude that these conformational dynamics were derived from the mobility of the C-terminal domain.

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Three-dimensional structure of calmodulin

Cited by 1,049 publications

References 42 publications

Kinetic Control of the Dissociation Pathway of Calmodulin-Peptide Complexes

Kinetic Control of the Dissociation Pathway of Calmodulin-Peptide Complexes

A series of point mutations reveal interactions between the calcium‐binding sites of calmodulin

Conformational dynamics of yeast calmodulin in the Ca²⁺‐bound state probed using NMR relaxation dispersion

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Three-dimensional structure of calmodulin

Cited by 1,049 publications

References 42 publications

Kinetic Control of the Dissociation Pathway of Calmodulin-Peptide Complexes

Kinetic Control of the Dissociation Pathway of Calmodulin-Peptide Complexes

A series of point mutations reveal interactions between the calcium‐binding sites of calmodulin

Conformational dynamics of yeast calmodulin in the Ca2+‐bound state probed using NMR relaxation dispersion

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Product

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Conformational dynamics of yeast calmodulin in the Ca²⁺‐bound state probed using NMR relaxation dispersion