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Molecular dynamics studies on troponin (TnI-TnT-TnC) complexes: insight into the regulation of muscle contraction.

ABSTRACT: Mutations of any subunit of the troponin complex may lead to serious disorders. Rational approaches to managing these disorders require knowledge of the complex interactions among the three subunits that are required for proper function. Molecular dynamics (MD) simulations were performed for both skeletal (sTn) and cardiac (cTn) troponin. The interactions and correlated motions among the three components of the troponin complex were analyzed using both Molecular Mechanics-Generalized Born Surface Area (MMGBSA) and cross-correlation techniques. The TnTH2 helix was strongly positively correlated with the two long helices of TnI. The C domain of TnC was positively correlated with TnI and TnT. The N domain of TnC was negatively correlated with TnI and TnT in cTn, but not in sTn. The two C-domain calcium-binding sites of TnC were dynamically correlated. The two regulatory N-domain calcium-binding sites of TnC were dynamically correlated, even though the calcium-binding site I is dysfunctional. The strong interaction residue pairs and the strong dynamically correlated residues pairs among the three components of troponin complexes were identified. These correlated motions are consistent with the idea that there is a high degree of cooperativity among the components of the regulatory complex in response to Ca(2+) and other effectors. This approach may give insight into the mechanism by which mutations of troponin cause disease. It is interesting that some observed disease causing mutations fall within regions of troponin that are strongly correlated or interacted.

SUBMITTER: Varughese JF

PROVIDER: S-EPMC3807689 | biostudies-literature | 2010 Oct

REPOSITORIES: biostudies-literature

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Molecular dynamics studies on troponin (TnI-TnT-TnC) complexes: insight into the regulation of muscle contraction.

Varughese Jayson F JF Chalovich Joseph M JM Li Yumin Y

Journal of biomolecular structure & dynamics 20101001 2

Mutations of any subunit of the troponin complex may lead to serious disorders. Rational approaches to managing these disorders require knowledge of the complex interactions among the three subunits that are required for proper function. Molecular dynamics (MD) simulations were performed for both skeletal (sTn) and cardiac (cTn) troponin. The interactions and correlated motions among the three components of the troponin complex were analyzed using both Molecular Mechanics-Generalized Born Surfac ...[more]

PMID: 20645650

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Project description:The heterotrimeric cardiac troponin complex is a key regulator of contraction and plays an essential role in conferring Ca2+ sensitivity to the sarcomere. During ischemic injury, rapidly accumulating protons acidify the myoplasm, resulting in markedly reduced Ca2+ sensitivity of the sarcomere. Unlike the adult heart, sarcomeric Ca2+ sensitivity in fetal cardiac tissue is comparatively pH insensitive. Replacement of the adult cardiac troponin I (cTnI) isoform with the fetal troponin I (ssTnI) isoform renders adult cardiac contractile machinery relatively insensitive to acidification. Alignment and functional studies have determined histidine 132 of ssTnI to be the predominant source of this pH insensitivity. Substitution of histidine at the cognate position 164 in cTnI confers the same pH insensitivity to adult cardiac myocytes. An alanine at position 164 of cTnI is conserved in all mammals, with the exception of the platypus, which expresses a proline. Prolines are biophysically unique because of their innate conformational rigidity and helix-disrupting function. To provide deeper structure-function insight into the role of the TnC-TnI interface in determining contractility, we employed a live-cell approach alongside molecular dynamics simulations to ascertain the chemo-mechanical implications of the disrupted helix 4 of cTnI where position 164 exists. This important motif belongs to the critical switch region of cTnI. Substitution of a proline at position 164 of cTnI in adult rat cardiac myocytes causes increased contractility independent of alterations in the Ca2+ transient. Free-energy perturbation calculations of cTnC-Ca2+ binding indicate no difference in cTnC-Ca2+ affinity. Rather, we propose the enhanced contractility is derived from new salt bridge interactions between cTnI helix 4 and cTnC helix A, which are critical in determining pH sensitivity and contractility. Molecular dynamics simulations demonstrate that cTnI A164P structurally phenocopies ssTnI under baseline but not acidotic conditions. These findings highlight the evolutionarily directed role of the TnI-cTnC interface in determining cardiac contractility.

Project description:Small molecule cardiac troponin activators could potentially enhance cardiac muscle contraction in the treatment of systolic heart failure. We designed a small molecule, RPI-194, to bind cardiac/slow skeletal muscle troponin (Cardiac muscle and slow skeletal muscle share a common isoform of the troponin C subunit.) Using solution NMR and stopped flow fluorescence spectroscopy, we determined that RPI-194 binds to cardiac troponin with a dissociation constant KD of 6-24 μM, stabilizing the activated complex between troponin C and the switch region of troponin I. The interaction between RPI-194 and troponin C is weak (KD 311 μM) in the absence of the switch region. RPI-194 acts as a calcium sensitizer, shifting the pCa50 of isometric contraction from 6.28 to 6.99 in mouse slow skeletal muscle fibers and from 5.68 to 5.96 in skinned cardiac trabeculae at 100 μM concentration. There is also some cross-reactivity with fast skeletal muscle fibers (pCa50 increases from 6.27 to 6.52). In the slack test performed on the same skinned skeletal muscle fibers, RPI-194 slowed the velocity of unloaded shortening at saturating calcium concentrations, suggesting that it slows the rate of actin-myosin cross-bridge cycling under these conditions. However, RPI-194 had no effect on the ATPase activity of purified actin-myosin. In isolated unloaded mouse cardiomyocytes, RPI-194 markedly decreased the velocity and amplitude of contractions. In contrast, cardiac function was preserved in mouse isolated perfused working hearts. In summary, the novel troponin activator RPI-194 acts as a calcium sensitizer in all striated muscle types. Surprisingly, it also slows the velocity of unloaded contraction, but the cause and significance of this is uncertain at this time. RPI-194 represents a new class of non-specific troponin activator that could potentially be used either to enhance cardiac muscle contractility in the setting of systolic heart failure or to enhance skeletal muscle contraction in neuromuscular disorders.

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Molecular dynamics studies on troponin (TnI-TnT-TnC) complexes: insight into the regulation of muscle contraction.

Publications

Molecular dynamics studies on troponin (TnI-TnT-TnC) complexes: insight into the regulation of muscle contraction.

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