title: Atomic resolution probe for allostery in the regulatory thin filament.
abstract: Calcium binding and dissociation within the cardiac thin filament (CTF) is a fundamental regulator of normal contraction and relaxation. Although the disruption of this complex, allosterically mediated process has long been implicated in human disease, the precise atomic-level mechanisms remain opaque, greatly hampering the development of novel targeted therapies. To address this question, we used a fully atomistic CTF model to test both Ca(2+) binding strength and the energy required to remove Ca(2+) from the N-lobe binding site in WT and mutant troponin complexes that have been linked to genetic cardiomyopathies. This computational approach is combined with measurements of in vitro Ca(2+) dissociation rates in fully reconstituted WT and cardiac troponin T R92L and R92W thin filaments. These human disease mutations represent known substitutions at the same residue, reside at a significant distance from the calcium binding site in cardiac troponin C, and do not affect either the binding pocket affinity or EF-hand structure of the binding domain. Both have been shown to have significantly different effects on cardiac function in vivo. We now show that these mutations independently alter the interaction between the Ca(2+) ion and cardiac troponin I subunit. This interaction is a previously unidentified mechanism, in which mutations in one protein of a complex indirectly affect a third via structural and dynamic changes in a second to yield a pathogenic change in thin filament function that results in mutation-specific disease states. We can now provide atom-level insight that is potentially highly actionable in drug design.
authors.affiliation: Department of Chemistry and Biochemistry, The University of Arizona, Tucson, AZ 85721;
authors.forename: Michael R
authors.initials: MR
authors.lastname: Williams
authors.affiliation: Physiological Sciences, The University of Arizona, Tucson, AZ 85724;
authors.forename: Sarah J
authors.initials: SJ
authors.lastname: Lehman
authors.affiliation: Department of Medicine, The University of Arizona, Tucson, AZ 85724; Department of Cellular and Molecular Medicine, The University of Arizona, Tucson, AZ 85724.
authors.forename: Jil C
authors.initials: JC
authors.lastname: Tardiff
authors.affiliation: Department of Chemistry and Biochemistry, The University of Arizona, Tucson, AZ 85721; sschwartz@email.arizona.edu.
authors.forename: Steven D
authors.initials: SD
authors.lastname: Schwartz
mesh_terms: D000494:Allosteric Regulation; D002118:Calcium / Q000378:metabolism; D006801:Humans; D008956:Models, Chemical; D020107:Troponin T / Q000737:chemistry / Q000378:metabolism*
chemical_list: D020107:Troponin T; D002118:Calcium
keywords: calcium homeostasis; cardiac thin filament; hypertrophic cardiomyopathy; molecular modeling; steered molecular dynamics