TnI Structural Interface with the N-Terminal Lobe of TnC as a Determinant of Cardiac Contractility

Abstract

The heterotrimeric cardiac troponin complex is a key regulator of contraction and plays an essential role in conferring Ca sensitivity to the sarcomere. During ischemic injury, rapidly accumulating protons acidify the myoplasm, resulting in markedly reduced Ca sensitivity of the sarcomere. Unlike the adult heart, sarcomeric Ca 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 Ca transient. Free-energy perturbation calculations of cTnC-Ca binding indicate no difference in cTnC-Ca 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.

Computational Studies of Cardiac and Skeletal Troponin

“…These simulations were able to calculate the work required to pull calcium ions from cNTnC within the context of the core of cTn. Free energy perturbations from the Metzger group were able to show an increase in calcium binding free energy for acidosis states of myocytes that agreed with experimental data (Thompson et al, 2014; Vetter et al, 2018). Finally, a four state model was developed to explain, through investigation of the cTnI effective concentration, why calcium sensitivity varies from isolated TnC to the Tn complex to a full thin filament model (Siddiqui et al, 2016).…”

“…Continuing their investigation into the role of isoform-specific interactions, the Metzger group subsequently simulated cTnC-cTnI switch-peptide systems at various protonation states (Palpant et al, 2012). Further study of the cTnC-cTnI in complex showed that there are key structural differences between the helix 4 of TnI and the switch-peptide region (Vetter et al, 2018). The intrinsically disordered region of TnI (C-terminal domain) was simulated with cNTnC which provided evidence that the region is flexible and has structural preferences (Metskas and Rhoades, 2015).…”

Levosimendan increases the phosphorylation state of phospholamban in the isolated human atrium

“…We assume this is probably due to a surge in the phosphorylation state of phospholamban and of the phosphorylation state of the inhibitory subunit of troponin that we measured here. Cardiac relaxation is renowned to be enhanced by phosphorylation of phospholamban (Hamstra et al 2020 ) and phosphorylation of the inhibitory subunit of troponin (Vetter et al 2018 ). Positive inotropic effects of levosimendan in human atrial preparations have been delineated before by others (Sahin et al 2007 ; Usta et al 2004 ).…”

Prognostic value of prealbumin, N-terminal pro-B-type natriuretic peptide, heart type fatty acid binding protein, and cardiac troponin I in elderly patients for heart failure and poor outcomes

“… 15 Cardiac troponin I (cTnI) is the molecular switch of the sarcomere and is related to relaxation performance in cardiac myocytes. 16 , 17 …”

“…15 Cardiac troponin I (cTnI) is the molecular switch of the sarcomere and is related to relaxation performance in cardiac myocytes. 16,17 In this study, we evaluated the combined prognostic value of prealbumin, NT-proBNP, hFABP, and cTnI for HF in elderly patients who are hospitalized because of chronic HF. Our findings could help evaluate and stratify the severity and risk of death in patients with HF and provide monitoring tools for treatment of this disease.…”