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Molecular dynamics simulations of the cardiac troponin complex performed with FRET distances as restraints.

Jayasundar JJ, Xing J, Robinson JM, Cheung HC, Dong WJ - PLoS ONE (2014)

Bottom Line: In the presence of saturating Ca(2+) the above said phenomenon were absent.We postulate that the secondary structure perturbations experienced by the cTnI regulatory region held within the cTnC N-domain hydrophobic pocket, coupled with the rotation of the cTnC N-domain would control the cTnI mobile domain interaction with actin.Concomitantly the rotation of the cTnC N-domain and perturbation of the D/E linker rigidity would control the cTnI inhibitory region interaction with actin to effect muscle relaxation.

View Article: PubMed Central - PubMed

Affiliation: Voiland School of Chemical Engineering and Bioengineering and The Department of Integrated Physiology and Neuroscience, Washington State University, Pullman, Washington, United States of America.

ABSTRACT
Cardiac troponin (cTn) is the Ca(2+)-sensitive molecular switch that controls cardiac muscle activation and relaxation. However, the molecular detail of the switching mechanism and how the Ca(2+) signal received at cardiac troponin C (cTnC) is communicated to cardiac troponin I (cTnI) are still elusive. To unravel the structural details of troponin switching, we performed ensemble Förster resonance energy transfer (FRET) measurements and molecular dynamic (MD) simulations of the cardiac troponin core domain complex. The distance distributions of forty five inter-residue pairs were obtained under Ca(2+)-free and saturating Ca(2+) conditions from time-resolved FRET measurements. These distances were incorporated as restraints during the MD simulations of the cardiac troponin core domain. Compared to the Ca(2+)-saturated structure, the absence of regulatory Ca(2+) perturbed the cTnC N-domain hydrophobic pocket which assumed a closed conformation. This event partially unfolded the cTnI regulatory region/switch. The absence of Ca(2+), induced flexibility to the D/E linker and the cTnI inhibitory region, and rotated the cTnC N-domain with respect to rest of the troponin core domain. In the presence of saturating Ca(2+) the above said phenomenon were absent. We postulate that the secondary structure perturbations experienced by the cTnI regulatory region held within the cTnC N-domain hydrophobic pocket, coupled with the rotation of the cTnC N-domain would control the cTnI mobile domain interaction with actin. Concomitantly the rotation of the cTnC N-domain and perturbation of the D/E linker rigidity would control the cTnI inhibitory region interaction with actin to effect muscle relaxation.

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Electrostatic surface analysis of the cTnC and cTnI.Depicts the electrostatics in the vicinity of cTnC helix E in the Ca2+-free state after 9.5 ns of simulation. The hydrophobic, negative and positive surfaces are colored white, red and blue respectively. (a) The thick arrow points to the unfolded segment in helix E. The positively charged blue residues of cTnI-Ir are seen arching (pointed to by the dotted line with arrow head) towards the unfolded cTnC helix E (pointed to by thick black arrow). The amino acids sequence of the cTnI-Ir residues is 138-KFKRLPT and the sequence of the opposing cTnC residues are 92-KSEEEL. The predominantly negative (red) cTnC helix E is attracted to the positive region of cTnI-Ir. The unfolded helix E has adopted a “U” shape (pointed to by the thick black arrow). (b) The unfolded helix E is seen in concert with cTnI and cTnT. The cTnC Glu94 is attracted to cTnI Lys141 (not seen in picture), cTnC Glu95 is attracted to the nitrogen on Leu129 of cTnI and Arg142 of cTnI, and cTnC Glu96 is attracted to Arg 267 of cTnT.
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pone-0087135-g006: Electrostatic surface analysis of the cTnC and cTnI.Depicts the electrostatics in the vicinity of cTnC helix E in the Ca2+-free state after 9.5 ns of simulation. The hydrophobic, negative and positive surfaces are colored white, red and blue respectively. (a) The thick arrow points to the unfolded segment in helix E. The positively charged blue residues of cTnI-Ir are seen arching (pointed to by the dotted line with arrow head) towards the unfolded cTnC helix E (pointed to by thick black arrow). The amino acids sequence of the cTnI-Ir residues is 138-KFKRLPT and the sequence of the opposing cTnC residues are 92-KSEEEL. The predominantly negative (red) cTnC helix E is attracted to the positive region of cTnI-Ir. The unfolded helix E has adopted a “U” shape (pointed to by the thick black arrow). (b) The unfolded helix E is seen in concert with cTnI and cTnT. The cTnC Glu94 is attracted to cTnI Lys141 (not seen in picture), cTnC Glu95 is attracted to the nitrogen on Leu129 of cTnI and Arg142 of cTnI, and cTnC Glu96 is attracted to Arg 267 of cTnT.

Mentions: As we traverse towards the D/E linker region, in the Ca2+-free state the D/E linker residues N-terminus of Asp87 were positioned towards the cTnI-Ir. We postulate that the folding and unfolding of helix D residues 82–86 in response Ca2+ flux pulls and releases the D/E linker residues, which in turn translate to the unfolding and refolding of cTnC helix E. The unfolded helix E residues (94–97) in the Ca2+-free state, were seen arching towards the cTnI-Ir (138–144) (Figure 6). We postulate that the unfolding of helix E probably helps control the release of the cTnI inhibitory region to interact with actin in the thin filament. Closer examination of the electrostatic surface of the troponin complex revealed that the unfolded region of cTnC is predominantly negative with three Glu residues. These negative residues were attracted to positively charged residues on TnI (Figure 6). It is known that the flexibility of the central helices of cTnC have a significant functional role in muscle regulation as cTnC mutants with altered central linker lengths or reduced linker flexibility are ineffective in the regulation of actomyosin ATPase [30]–[32]. The observed Ca2+-induced changes in the structural dynamics of the central helices of cTnC may provide molecular basis of Ca2+ signaling in the cardiac thin filament regulation. We postulate that the Ca2+ flux based folding and unfolding of helix D of cTnC propagates the Ca2+ signal to cTnI-Ir via the D/E linker of cTnC, which adopts a flexible conformation in the Ca2+-free state and adopts a rigid conformation in the Ca2+-saturated state. The alternating flexibility of the D/E linker regions probably releases and retracts the cTnI-Ir, towards or away from actin.


Molecular dynamics simulations of the cardiac troponin complex performed with FRET distances as restraints.

Jayasundar JJ, Xing J, Robinson JM, Cheung HC, Dong WJ - PLoS ONE (2014)

Electrostatic surface analysis of the cTnC and cTnI.Depicts the electrostatics in the vicinity of cTnC helix E in the Ca2+-free state after 9.5 ns of simulation. The hydrophobic, negative and positive surfaces are colored white, red and blue respectively. (a) The thick arrow points to the unfolded segment in helix E. The positively charged blue residues of cTnI-Ir are seen arching (pointed to by the dotted line with arrow head) towards the unfolded cTnC helix E (pointed to by thick black arrow). The amino acids sequence of the cTnI-Ir residues is 138-KFKRLPT and the sequence of the opposing cTnC residues are 92-KSEEEL. The predominantly negative (red) cTnC helix E is attracted to the positive region of cTnI-Ir. The unfolded helix E has adopted a “U” shape (pointed to by the thick black arrow). (b) The unfolded helix E is seen in concert with cTnI and cTnT. The cTnC Glu94 is attracted to cTnI Lys141 (not seen in picture), cTnC Glu95 is attracted to the nitrogen on Leu129 of cTnI and Arg142 of cTnI, and cTnC Glu96 is attracted to Arg 267 of cTnT.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3928104&req=5

pone-0087135-g006: Electrostatic surface analysis of the cTnC and cTnI.Depicts the electrostatics in the vicinity of cTnC helix E in the Ca2+-free state after 9.5 ns of simulation. The hydrophobic, negative and positive surfaces are colored white, red and blue respectively. (a) The thick arrow points to the unfolded segment in helix E. The positively charged blue residues of cTnI-Ir are seen arching (pointed to by the dotted line with arrow head) towards the unfolded cTnC helix E (pointed to by thick black arrow). The amino acids sequence of the cTnI-Ir residues is 138-KFKRLPT and the sequence of the opposing cTnC residues are 92-KSEEEL. The predominantly negative (red) cTnC helix E is attracted to the positive region of cTnI-Ir. The unfolded helix E has adopted a “U” shape (pointed to by the thick black arrow). (b) The unfolded helix E is seen in concert with cTnI and cTnT. The cTnC Glu94 is attracted to cTnI Lys141 (not seen in picture), cTnC Glu95 is attracted to the nitrogen on Leu129 of cTnI and Arg142 of cTnI, and cTnC Glu96 is attracted to Arg 267 of cTnT.
Mentions: As we traverse towards the D/E linker region, in the Ca2+-free state the D/E linker residues N-terminus of Asp87 were positioned towards the cTnI-Ir. We postulate that the folding and unfolding of helix D residues 82–86 in response Ca2+ flux pulls and releases the D/E linker residues, which in turn translate to the unfolding and refolding of cTnC helix E. The unfolded helix E residues (94–97) in the Ca2+-free state, were seen arching towards the cTnI-Ir (138–144) (Figure 6). We postulate that the unfolding of helix E probably helps control the release of the cTnI inhibitory region to interact with actin in the thin filament. Closer examination of the electrostatic surface of the troponin complex revealed that the unfolded region of cTnC is predominantly negative with three Glu residues. These negative residues were attracted to positively charged residues on TnI (Figure 6). It is known that the flexibility of the central helices of cTnC have a significant functional role in muscle regulation as cTnC mutants with altered central linker lengths or reduced linker flexibility are ineffective in the regulation of actomyosin ATPase [30]–[32]. The observed Ca2+-induced changes in the structural dynamics of the central helices of cTnC may provide molecular basis of Ca2+ signaling in the cardiac thin filament regulation. We postulate that the Ca2+ flux based folding and unfolding of helix D of cTnC propagates the Ca2+ signal to cTnI-Ir via the D/E linker of cTnC, which adopts a flexible conformation in the Ca2+-free state and adopts a rigid conformation in the Ca2+-saturated state. The alternating flexibility of the D/E linker regions probably releases and retracts the cTnI-Ir, towards or away from actin.

Bottom Line: In the presence of saturating Ca(2+) the above said phenomenon were absent.We postulate that the secondary structure perturbations experienced by the cTnI regulatory region held within the cTnC N-domain hydrophobic pocket, coupled with the rotation of the cTnC N-domain would control the cTnI mobile domain interaction with actin.Concomitantly the rotation of the cTnC N-domain and perturbation of the D/E linker rigidity would control the cTnI inhibitory region interaction with actin to effect muscle relaxation.

View Article: PubMed Central - PubMed

Affiliation: Voiland School of Chemical Engineering and Bioengineering and The Department of Integrated Physiology and Neuroscience, Washington State University, Pullman, Washington, United States of America.

ABSTRACT
Cardiac troponin (cTn) is the Ca(2+)-sensitive molecular switch that controls cardiac muscle activation and relaxation. However, the molecular detail of the switching mechanism and how the Ca(2+) signal received at cardiac troponin C (cTnC) is communicated to cardiac troponin I (cTnI) are still elusive. To unravel the structural details of troponin switching, we performed ensemble Förster resonance energy transfer (FRET) measurements and molecular dynamic (MD) simulations of the cardiac troponin core domain complex. The distance distributions of forty five inter-residue pairs were obtained under Ca(2+)-free and saturating Ca(2+) conditions from time-resolved FRET measurements. These distances were incorporated as restraints during the MD simulations of the cardiac troponin core domain. Compared to the Ca(2+)-saturated structure, the absence of regulatory Ca(2+) perturbed the cTnC N-domain hydrophobic pocket which assumed a closed conformation. This event partially unfolded the cTnI regulatory region/switch. The absence of Ca(2+), induced flexibility to the D/E linker and the cTnI inhibitory region, and rotated the cTnC N-domain with respect to rest of the troponin core domain. In the presence of saturating Ca(2+) the above said phenomenon were absent. We postulate that the secondary structure perturbations experienced by the cTnI regulatory region held within the cTnC N-domain hydrophobic pocket, coupled with the rotation of the cTnC N-domain would control the cTnI mobile domain interaction with actin. Concomitantly the rotation of the cTnC N-domain and perturbation of the D/E linker rigidity would control the cTnI inhibitory region interaction with actin to effect muscle relaxation.

Show MeSH
Related in: MedlinePlus